bluuebunny/crossref_metadata_2025 huggingface dataset:
A polymorphic CA repeat in the IGF‐I gene is associated with gender‐specific differences in body height, but has no effect on the secular trend in body height
Authors: Rietveld I. , Janssen J. A. M. J. L. , Van Rossum E. F. C. , Houwing‐Duistermaat J. J. , Rivadeneira F. , Hofman A. , Pols H. A. P. , Van Duijn C. M. , Lamberts S. W. J.
Published: July 2004

objective  A polymorphism near the promoter region of the IGF‐I gene has been associated with serum IGF‐I levels, age‐related decline of serum IGF‐I levels, body height, birth weight and intima media thickness in hypertensive subjects.

design and methods  We investigated the association between the length of the IGF‐I alleles of this promoter polymorphism and IGF‐I levels and body height. Furthermore, we investigated the potential influence of this polymorphism on final height in relationship to the secular trend of individuals born between 1917 and 1945. All subjects were participants of the Rotterdam Study.

results  We observed, in analyses including only homozygous carriers, the highest IGF‐I levels in homozygous carriers of the 192‐bp allele (18·7 nmol/l ± 0·6) and homozygous carriers of the 194‐bp allele (17·7 nmol/l ± 1·4). IGF‐I levels were significantly lower in individuals with homozygous longer alleles [> 194‐bp (12·0 nmol/l ± 1·2; P < 0·001)] and homozygous shorter alleles [< 192‐bp (15·6 nmol/l ± 1·4; P < 0·05)] compared to homozygous carriers of the 192‐bp and the 194‐bp allele. In males and females separately, an optimum for serum IGF‐I was also observed in homozygous carriers of the 192‐bp and 194‐bp allele. Only in males, homozygous carriers of the 192‐bp allele were significantly taller than homozygous carriers of the shorter alleles (174·9 cm ± 0·2 vs. 171·5 cm ± 1·4; P = 0·01). When all subjects genotyped for the IGF‐I promoter polymorphism were included in the analysis, a clear optimum for IGF‐I levels and body height was observed in carriers of the 192‐bp and/or 194‐bp allele in the total population. Between 1917 and 1945, a secular trend in body height was observed in our Dutch population. Mean final body height was significantly higher in carriers of the most frequent alleles (192‐bp and/or the 194‐bp), than carriers of the remaining shorter and longer genotypes (P‐trend < 0·01).

conclusions  In conclusion, we observed an optimum in IGF‐I levels and final body height for the 192‐bp and 194‐bp allele of the IGF‐I gene. A gender‐specific effect of the IGF‐I alleles on body height was observed. The secular trend in body height observed in our elderly Dutch population was similar for the different genotypes; carriers of the 192‐bp and/or the 194‐bp allele remained significantly taller throughout time.
Lack of independent relationship between plasma adiponectin, leptin levels and bone density in nondiabetic female adolescents
Authors: Huang Kuo‐Chin , Cheng Wern‐Cherng , Yen Ruoh‐Fang , Tsai Keh‐Sung , Tai Tong‐Yuan , Yang Wei‐Shiung
Published: July 2004

objectives  Adiponectin has been implicated in the pathophysiology of metabolic syndrome and coronary artery disease in humans. Whether adiponectin is related to bone mineralization remains unclear in adults as well as in adolescents. In this study, we aimed to determine the relationship between plasma adiponectin, leptin concentrations and bone density, including total‐body bone mineral density (BMD) and bone mineral content (BMC) in adolescence.

patients and measurements  We studied 105 nondiabetic female adolescents [mean age 15·4 ± 1·9 years, and mean body mass index (BMI), 23·1 ± 4·0 kg/m2]. A venous blood sample was taken after 12 h of fasting to measure fasting plasma adiponectin and leptin levels. BMD and BMC of the whole body were measured by dual‐energy X‐ray absorptiometry.

results  In simple correlation analysis, plasma adiponectin and leptin levels correlated significantly with total‐body BMD (r =−0·523 and r = 0·443, P < 0·001, respectively) and BMC (r =−0·471 and r = 0·396, P < 0·001, respectively). However, plasma adiponectin and leptin were related to both BMD and BMC in opposite directions. In multivariate linear regression analyses, only BMI or fat mass (FM) and Tanner stage, but not plasma adiponectin and leptin, were significantly related to BMD and BMC following adjustment for other variables.

conclusions  The results suggest that plasma adiponectin and leptin concentrations are not related to the total‐body BMD and BMC independent of the chronological age, BMI or FM, and Tanner stage in nondiabetic female adolescents, although they were highly correlated in simple correlation analyses. The biologic roles of adiponectin in bone still need further clarification.
Cabergoline addition to depot somatostatin analogues in resistant acromegalic patients: efficacy and lack of predictive value of prolactin status
Authors: Cozzi Renato , Attanasio Roberto , Lodrini Sandro , Lasio Giovanni
Published: July 2004

background  Somatostatin analogues (SA) are currently the mainstay in the medical treatment of acromegaly. However, even high doses of depot SA for prolonged periods do not achieve GH–IGF‐I normalization in some patients. Even though some data were reported about the addition of cabergoline, a long‐acting dopamine agonist (DA), to SA in resistant patients, definite data are still lacking.

design  Prospective open trial.

patients  In 19 acromegalic patients with active disease (34–82 years old, seven males, 12 females) resistant to chronic (9–12 months) depot SA (octreotide‐LAR, 30 mg/28 days in 13 patients, lanreotide, 60 mg/28 days intramuscularly in six patients) cabergoline was added (combined treatment). In these patients, SA treatment had partially relieved GH and IGF‐I hypersecretion but no patient had achieved ‘safe’ GH and normal IGF‐I‐values. Eight patients had PRL levels greater than 15 µg/l (range 16–60 µg/l; 1 µg = 21·2 mIU). Immunohistochemistry (IHC) was positive for PRL in four out of eight operated patients.

results  The addition of cabergoline, using the minimal effective and the maximal tolerated dose (range 1–3·5 mg/week), decreased GH from 6·6 ± 0·9 to 4·6 ± 0·6 µg/l (P = 0·018), and IGF‐I from 552 ± 44 to 428 ± 54 µg/l (P = 0·019) after 6 months (median, range 3–18 Months). Combined treatment decreased GH to < 2·5 µg/l in four patients (21%) and normalized IGF‐I for age in eight patients (42%). It obtained a decline of both GH and IGF‐I (−49 ± 7%, and −47 ± 5%, respectively) in nine patients (47%), and a partial improvement in six (32%) patients (GH decreased by 43 ± 8% in four, and IGF‐I by 35–38% in two patients). No change was observed in two patients, and worsening in two other patients. Results were not dependent on PRL status (serum levels or IHC). Combined treatment was well tolerated.

conclusions  The addition of cabergoline to depot SA‐resistant acromegalic patients is effective, not dependent on PRL values and normalizes IGF‐I levels in 42% of patients. The association of long‐acting DA and SA deserves a more relevant role in the therapeutical algorithm of acromegaly.
Polymorphisms in oestrogen and progesterone receptor genes: possible influence on prolactin levels in women
Authors: Westberg Lars , Ho Hoi‐Por , Baghaei Fariba , Nilsson Staffan , Melke Jonas , Rosmond Roland , Holm Göran , Björntorp Per , Eriksson Elias
Published: July 2004

objective  Oestrogen and progesterone are known to influence the release of human prolactin. The present study was undertaken in order to investigate the possible influence of polymorphisms of the genes encoding the oestrogen receptor (ER)α, ERβ and the progesterone receptor (PGR), on prolactin levels in premenopausal women.

design and measurements  Serum levels of prolactin were measured in the follicular phase of the menstrual cycle. Subjects were genotyped with respect to a TA repeat polymorphism of the ERα gene, a CA repeat polymorphism of the ERβ gene, and two polymorphisms of the PGR gene: one insertion polymorphism (PROGINS) and one single nucleotide polymorphism (G331A).

subjects  A population‐based cohort of 270 42‐year‐old women.

results  The CA repeat polymorphism of the ERβ gene and the G331A polymorphism of the PGR gene appeared to be associated with prolactin levels. In contrast, we found no evidence for an influence of the PROGINS polymorphism of the PGR gene or the TA repeat polymorphism of the ERα gene on the levels of this hormone.

conclusions  These data suggest that genetic variants of both the ERβ and the PGR may influence prolactin release.
Systematic dose‐extension of octreotide LAR: the importance of individual tailoring of treatment in patients with acromegaly
Authors: Turner Helen E. , Thornton‐Jones Viv A. , Wass John A. H.
Published: July 2004

objective  The depot long‐acting somatostatin analogue octreotide LAR (LAR) provides effective and well‐tolerated treatment for acromegaly. Despite a 4‐weekly recommended injection frequency, prolonged duration of GH suppression has been observed in some patients following treatment with long‐acting somatostatin analogues. The aim of our study was to perform a prospective systematic study to determine whether extending the interval between doses of LAR allows maintenance of ‘safe’ GH in selected patients with acromegaly.

patients and methods  Twenty‐two patients (15 men, seven women), mean age 58·9 years (35–81 years) with active acromegaly (mGH > 5 mU/l), requiring treatment were selected to receive treatment with LAR. Eleven patients had received previous treatment with both transsphenoidal surgery and radiotherapy, while six had received surgery alone. All patients were commenced on treatment with 20 mg LAR intramuscularly (i.m.) every 4 weeks. Mean GH (mGH) was measured after three consecutive injections immediately prior to the fourth injection. The dose frequency was systematically reduced after every four injections if mGH < 5 mU/l. Once mGH > 5 mU/l, the dose frequency was increased and mGH reassessed.

results  The dosing interval was successfully increased to greater than 4 weeks in 20/22 patients (90·9%). Six of 22 (27·3%) were receiving injections every 8 weeks and 3/22 (13·6%) every 12 weeks. GH and IGF‐I were lower on treatment compared with baseline (P < 0·01). There was no difference in individual mGH and IGF‐I between the values on 4‐weekly dosing and those at final dose frequency. There was no relationship between final dose frequency and either mean GH or IGF‐I prior to LAR, patient age, or previous treatment. The percentage suppression following 100 µg octreotide subcutaneously did not predict subsequent dose frequency of LAR. The drug cost if patients had continued at 4‐weekly intervals would be UK£187 850, compared with UK£101 065 for the individually titrated dose frequency (P < 0·01). This represents a final cost of 53·8% of the 4‐weekly injection price.

conclusion  Individual tailoring of LAR administration maintains control of acromegaly, with reduced injection frequency and improved cost‐effectiveness.
Abnormal regulation of thirst and vasopressin secretion following surgery for craniopharyngioma
Authors: Smith D. , Finucane F. , Phillips J. , Baylis P. H. , Finucane J. , Tormey W. , Thompson C. J.
Published: July 2004

objective  In this study we aimed to establish the frequency of postoperative diabetes insipidus and the incidence and characteristics of abnormalities of thirst in a cohort of patients with craniopharyngioma, in whom neurosurgery had been performed.

design  Diabetes insipidus was determined by either standard criteria for diagnosis in the immediate postoperative period, or by water deprivation test, in all craniopharyngioma and pituitary tumour patients who underwent surgery in Beaumont Hospital between the years 1986 and 1998. Osmoregulated thirst and vasopressin release were studied during a 2‐h infusion of hypertonic (5%) saline followed by a 30‐min period of free access to water.

patients  Data on the incidence of postoperative diabetes insipidus was collected in 26 patients with craniopharyngioma and 154 patients with pituitary adenomata. We recruited 16 healthy control patients, 16 patients with cranial diabetes insipidus following pituitary tumour surgery and 16 patients with cranial diabetes insipidus following craniopharyngioma resection for the hypertonic saline infusion study.

results  Twenty‐five patients out of 26 (96%) patients developed diabetes insipidus after surgery for craniopharyngioma, a much higher incidence than after surgery for suprasellar (26/88, 30%, P < 0·001) or intrasellar pituitary tumours (9/66, 14%, P < 0·001). Hypertonic saline infusion identified abnormal thirst responses in five of the 16 craniopharygioma patients studied; all of the pituitary tumour patients had a normal thirst response. Three of the craniopharyngioma patients had adipsic diabetes insipidus whilst two had polydipsic diabetes insipidus.

conclusion  This study demonstrates following surgery for craniopharyngioma there is a high incidence of cranial diabetes insipidus and a significant incidence of abnormal thirst responses to osmotic stimuli.
Subclinical hypothyroidism is associated with a low‐grade inflammation, increased triglyceride levels and predicts cardiovascular disease in males below 50 years
Authors: Kvetny J. , Heldgaard P. E. , Bladbjerg E. M. , Gram J.
Published: July 2004

objective  Mild thyroid failure is associated with an increased risk for development of atherosclerosis, but whether subclinical hypothyroidism is related to risk for cardiovascular disease is controversial. The purpose of the present study was to examine a possible association between subclinical hypothyroidism and cardiovascular disease.

design  Cross‐sectional study of a general population.

patients  Twelve hundred and twelve subjects, men and women, between 20 and 69 years old without thyroid disease not treated with drugs interfering with thyroid function or analysis of TSH were included.

measurements  Clinical signs of cardiovascular disease based on a questionnaire and medical records and laboratory analysis of lipids, atherothrombotic risk markers, C‐reactive protein and TSH.

results  The main findings were a high incidence of subclinical hypothyroidism (19·7%) in a general population. Subclinical hypothyroidism was associated with higher concentrations of triglycerides and C‐reactive protein. Below 50 years of age cardiovascular disease was more frequent in males with subclinical hypothyroidism compared to euthyroid males. Subclinical hypothyroidism was a predictor of cardiovascular disease in males below 50 years with an odds ratio of 3·4 (95% confidence interval 1·6–6·8) for developing cardiovascular disease compared to euthyroid age‐matched males.

conclusion  Our study demonstrates that patients with subclinical hypothyroidism have increased levels of triglycerides and signs of low‐grade inflammation (raised C‐reactive protein levels) and that subclinical hypothyroidism might be a risk factor for development of cardiovascular disease in younger males.
BRAF mutations in an Italian cohort of thyroid cancers
Authors: Fugazzola Laura , Mannavola Deborah , Cirello Valentina , Vannucchi Guia , Muzza Marina , Vicentini Leonardo , Beck‐Peccoz Paolo
Published: July 2004

objective  Recently, a somatic point mutation of the BRAF gene (V599E) has been identified as the most common genetic event in papillary thyroid carcinoma (PTC) with a variable frequency (about 25–70%) in different series from USA, Japan, Portugal and Ukraine.

design  In the present study, the genetic analysis of BRAF in an Italian cohort of 65 thyroid tumours with corresponding normal tissues and 21 thyroid benign disorders is reported.

methods  For BRAF analysis, the somatic DNA was PCR amplified by means of specific intronic primers and PCR products were directly sequenced. Statistical analyses were obtained by means of Fisher's exact test.

results  All mutations detected involved a T > A transversion at 1796 (V599E) and were heterozygous. Overall, BRAFV599E mutation was found in 18/56 (32·1%) PTCs. According to the histological type of the tumour, the mutation was present in 38·3% of cases of conventional PTC (18/47), in 0/6 follicular variant of PTC, in 0/3 oncocytic variant of PTC. No BRAF mutations were detected either in five follicular carcinomas, or in four poorly differentiated or undifferentiated cancers or in benign thyroid disorders. No statistically significant correlation of BRAF mutation with patient age and gender, with multicentricity of the tumour, with the lymphocytic infiltration of the tissue, with the stage and with the recurrence rate, was found. BRAFV599E tended to be associated, although not significantly, with a greater volume and extension of the tumour and with lymph‐nodal metastases at surgery.

conclusions  In conclusion, the present study on the first Italian series of thyroid cancers shows a frequency of 38·3% of BRAFV599E in the classical variant of PTC, confirming the key role of this mutation in promoting tumourigenesis.
The relationship between circulating osteoprotegerin levels and bone mineral metabolism in healthy women
Authors: Oh Ki Won , Rhee Eun Jung , Lee Won Young , Kim San Woo , Oh Eun Sook , Baek Ki Hyun , Kang Moo Il , Choi Moon Gi , Yoo Hyung Joon , Park Sung Woo
Published: July 2004

objective  Osteoprotegerin (OPG) is a recently identified cytokine that acts as a decoy receptor for the RANK ligand. Moreover, OPG has been shown to be an important inhibitor of osteoclastogenesis in animal models. However, the relationship between circulating OPG levels and female bone status in human populations is unclear. In this study we undertook to investigate the relationship between circulating OPG levels and bone mineral metabolism in healthy women.

patients and measurements  Our subjects were 287 women aged 37–73 years (mean age 51·5 years). The serum concentrations of OPG were determined by enzyme‐linked immunosorbent assay (ELISA). The biochemical markers of bone turnover and FSH were measured using standard methods. Bone mineral densities at the lumbar spine and femoral neck were measured by dual‐energy X‐ray absorptiometry.

results  Postmenopausal women had a significantly higher mean value of serum OPG than premenopausal women (1358·5 ± 32·5 pg/ml vs. 1228·8 ± 33·3 pg/ml, P < 0·01). Serum OPG levels were positively correlated with age (r = 0·169, P < 0·01), as were urine deoxypyridinoline levels (r = 0·133, P < 0·05) and serum FSH levels (r = 0·187, P < 0·01) in a bivariate correlation analyses. In a multiple regression analysis, only urine calcium excretion was identified as a significant predictor for serum OPG levels.

conclusions  Circulating OPG levels were found to be associated with urine calcium excretion and menopause in healthy women. Our observations suggest that circulating OPG levels reflect an antiresorptive activity in bone, and they are related to endogenous oestrogen levels.
Effects of clomiphene and raloxifene on gonadotrophin secretion in postmenopausal women: evidence of nongenomic action
Authors: Garas A. , Trypsianis G. , Kallitsaris A. , Milingos S. , Messinis I. E.
Published: July 2004

objective  To evaluate the effect of raloxifene (R) and clomiphene (Cl) on FSH and LH secretion in postmenopausal women.

design  Postmenopausal volunteer women participated in two experimental (Exp) procedures. In Group 1, the women received R (180 mg/day orally) for 30 days plus oestradiol (E2) through skin patches (100 g/24 h) from days 21 to 30 (R‐Exp). After a month's break the same women received Cl (150 mg/day orally) for 30 days plus E2 as above (Cl‐Exp). In Group 2, the women received E2 for 30 days plus R from days 21 to 30 (R‐Exp) and after a month's break they received E2 for 30 days plus Cl from days 21 to 30 (Cl‐Exp). Daily doses were as in Group 1. A GnRH test (100 g intravenously) was performed in all women on days 0, 10, 20 and 30 of each experiment.

patients  Sixteen healthy postmenopausal women were divided into two groups (eight women in each group).

measurements  The area under the curve (AUC) of ΔFSH and ΔLH response to GnRH (net increase above the basal value) was calculated.

results  In Group 1, basal levels of FSH and LH did not change significantly during the R‐Exp, while they decreased significantly in the Cl‐Exp (P < 0·001). The addition of E2 did not have any effect. The AUC of LH response to GnRH increased significantly in the R‐Exp (P < 0·05) and that of FSH in the Cl‐Exp (P < 0·05). In Group 2, basal levels of FSH and LH declined significantly during treatment with E2 in both the R‐Exp (P < 0·01) and the Cl‐Exp (P < 0·001). However, the addition of Cl (for 10 days) interrupted this decrease, while the addition of R stimulated FSH levels significantly (P < 0·05). E2 suppressed significantly the AUC of LH in both experiments (P < 0·05). The addition of Cl did not affect the AUC in response to GnRH, while the addition of R increased the AUC of both LH and FSH (P < 0·05).

conclusions  These results demonstrate for the first time that in contrast to Cl, R does not exert oestrogenic effects on basal gonadotrophin secretion. Although the antioestrogenic action of these drugs was evident only after pretreatment with E2, both R and Cl stimulated GnRH‐induced gonadotrophin secretion in oestrogen‐deprived women. It is hypothesized that these two compounds sensitize the pituitary to GnRH through mechanisms not involving the oestrogen receptor complex (nongenomic).
Association of a thyroglobulin gene polymorphism with Hashimoto's thyroiditis in the Japanese population
Authors: Ban Yoshiyuki , Tozaki Teruaki , Taniyama Matsuo , Tomita Motowo , Ban Yoshio
Published: July 2004

objective  The aetiology of the autoimmune thyroid diseases (AITDs), Graves’ disease (GD) and Hashimoto's thyroiditis is largely unknown. However, genetic susceptibility is believed to play a major role. Two whole genome scans from Japan and from the USA identified a locus on chromosome 8q24 which showed evidence for linkage with AITD and HT. Recent studies have demonstrated an association between a Tg polymorphisms and AITD, suggesting that Tg is the susceptibility gene on 8q24.

patients  We studied 308 Japanese AITD patients (194 GD and 114 HT patients) and 417 Japanese control subjects in association studies.

design  Case–control association studies were performed using D8S284 and D8S272, microsatellite markers located in the 8q24 region, Tgms1 and Tgms2, microsatellite markers in introns 10 and 27, respectively, of Tg, and a SNP in exon 33 of Tg.

results  No differences in allele frequencies were observed between AITD patients and controls for D8S284, D8S272 and Tgms1. Similarly, for Tgms2 and the exon 33 SNP no significant differences in allele frequency distribution were observed for all AITD patients. However, when analysing the HT patients alone we found a significant association between the 330 bp/352 bp genotype of Tgms2 and HT (HT = 16·7%, controls = 7·1%; corrected P‐value = 0·01, OR = 2·6).

conclusion  Our results confirm the previous reports of an association between the Tg gene and AITD and suggest that Tg is an AITD susceptibility gene.
A single nucleotide polymorphism in the CD40 gene on chromosome 20q (GD‐2) provides no evidence for susceptibility to Graves’ disease in UK Caucasians
Authors: Heward Joanne M. , Simmonds Matthew J. , Carr‐Smith Jackie , Foxall Helen , Franklyn Jayne A. , Gough Stephen C. L.
Published: July 2004

objective  A genome‐wide screen in Graves’ disease (GD) has shown linkage to chromosome 20q, designated GD‐2. The gene encoding CD40, which stimulates lymphocyte proliferation and differentiation, maps to this region, and a single nucleotide polymorphism (SNP) at position −1 of the Kozak sequence within the gene has been reported to be associated with GD. The aim of this study was to determine whether this SNP of the CD40 gene confers susceptibility to GD in UK Caucasians.

design  A large case–control cohort consisting of 800 patients with GD, and 785 control subjects with no history of autoimmune disease, was used to genotype this SNP by polymerase chain reaction restriction fragment length polymorphism.

results  Despite adequate power (> 99%) to detect an effect, if present (odds ratio of 1·5), no significant difference in allele or genotype frequency of the CD40 SNP was observed between patients with GD and control subjects (P = 0·087 and P = 0·145, respectively).

conclusions  These data suggest that this polymorphism of the CD40 gene is not associated with GD in the UK and is therefore not contributing to disease susceptibility in the chromosomal region designated GD‐2.
MotPS is the stator‐force generator for motility of alkaliphilic Bacillus, and its homologue is a second functional Mot in Bacillus subtilis
Authors: Ito Masahiro , Hicks David B. , Henkin Tina M. , Guffanti Arthur A. , Powers Benjamin D. , Zvi Lior , Uematsu Katsuyuki , Krulwich Terry A.
Published: August 2004

The stator‐force generator that drives Na+‐dependent motility in alkaliphilic Bacillus pseudofirmus OF4 is identified here as MotPS, MotAB‐like proteins with genes that are downstream of the ccpA gene, which encodes a major regulator of carbon metabolism. B. pseudofirmus OF4 was only motile at pH values above 8. Disruption of motPS resulted in a non‐motile phenotype, and motility was restored by transformation with a multicopy plasmid containing the motPS genes. Purified and reconstituted MotPS from B. pseudofirmus OF4 catalysed amiloride analogue‐sensitive Na+ translocation. In contrast to B. pseudofirmus, Bacillus subtilis contains both MotAB and MotPS systems. The role of the motPS genes from B. subtilis in several motility‐based behaviours was tested in isogenic strains with intact motAB and motPS loci, only one of the two mot systems or neither mot system. B. subtilis MotPS (BsMotPS) supported Na+‐stimulated motility, chemotaxis on soft agar surfaces and biofilm formation, especially after selection of an up‐motile variant. BsMotPS also supported motility in agar soft plugs immersed in liquid; motility was completely inhibited by an amiloride analogue. BsMotPS did not support surfactin‐dependent swarming on higher concentration agar surfaces. These results indicate that BsMotPS contributes to biofilm formation and motility on soft agar, but not to swarming, in laboratory strains of B. subtilis in which MotAB is the dominant stator‐force generator. BsMotPS could potentially be dominant for motility in B. subtilis variants that arise in particular niches.
Defective cell wall synthesis in Streptococcus pneumoniae R6 depleted for the essential PcsB putative murein hydrolase or the VicR (YycF) response regulator
Authors: Ng Wai‐Leung , Kazmierczak Krystyna M. , Winkler Malcolm E.
Published: August 2004

PcsB is a protein of unknown function(s) that influences the cell morphology of several pathogenic species of streptococcus. PcsB contains a CHAP (cysteine, histidine‐dependent amidohydrolase/peptidase) domain found in bacterial murein hydrolases; however, direct links between steps in cell wall biosynthesis and PcsB function(s) have not been demonstrated. We show here that pcsB is essential in the human respiratory pathogen, Streptococcus pneumoniae, that depletion of PcsB is bacteriostatic and that alanine substitutions in the conserved cysteine and histidine residues of the CHAP domain appear to be lethal. We stained wild‐type parent and mutant bacteria deficient in expression of PcsB with fluorescent vancomycin and DAPI to determine patterns of cell wall synthesis and nucleoid segregation respectively. The wild‐type parent strain exhibited ordered, simultaneous septal and equatorial cell wall synthesis. In contrast, reduced expression of PcsB resulted in formation of long chains of cells in which peptidoglycan synthesis occurred at nearly every division septum and cell equator. Severe depletion of PcsB led to abnormal, uncontrolled cell wall synthesis at misplaced septa and around large cells. Together, these physiological properties are consistent with a role for PcsB as a murein hydrolase that balances the extent of cell wall synthesis in S. pneumoniae. Finally, we show that the defects in morphology and cell wall synthesis that result from depletion of PcsB strongly resemble those caused by depletion of the essential VicRK two component regulatory system (TCS). This result and the essentiality of pcsB support the hypothesis that the essentiality of the  VicRK TCS results from its positive regulation of PcsB expression.
GGDEF and EAL domains inversely regulate cyclic di‐GMP levels and transition from sessility to motility
Authors: Simm Roger , Morr Michael , Kader Abdul , Nimtz Manfred , Römling Ute
Published: August 2004

Cyclic nucleotides represent second messenger molecules in all kingdoms of life. In bacteria, mass sequencing of genomes detected the highly abundant protein domains GGDEF and EAL. We show here that the GGDEF and EAL domains are involved in the turnover of cyclic‐di‐GMP (c‐di‐GMP) in vivo whereby the GGDEF domain stimulates c‐di‐GMP production and the EAL domain c‐di‐GMP degradation. Thus, most probably, GGDEF domains function as c‐di‐GMP cyclase and EAL domains as phosphdiesterase. We further show that, in the pathogenic organism Salmonella enterica serovar Typhimurium, the nosocomial pathogen Pseudomonas aeruginosa and the commensal species Escherichia coli, GGDEF and EAL domains mediate similar phenotypic changes related to the transition between sessility and motility. Thus, the data suggest that c‐di‐GMP is a novel global second messenger in bacteria the metabolism of which is controlled by GGDEF and EAL domain proteins.
MicroReview: The role of Plasmodium falciparum var genes in malaria in pregnancy
Authors: Rowe J. A. , Kyes S. A.
Published: August 2004

Sequestration of Plasmodium falciparum‐infected erythrocytes in the placenta is responsible for many of the harmful effects of malaria during pregnancy. Sequestration occurs as a result of parasite adhesion molecules expressed on the surface of infected erythrocytes binding to host receptors in the placenta such as chondroitin sulphate A (CSA). Identification of the parasite ligand(s) responsible for placental adhesion could lead to the development of a vaccine to induce antibodies  to  prevent  placental  sequestration.  Such a vaccine would reduce the maternal anaemia and infant deaths that are associated with malaria in pregnancy. Current research indicates that the parasite ligands mediating placental adhesion may be members of the P. falciparum variant surface antigen family PfEMP1, encoded by var genes. Two relatively well‐conserved subfamilies of var genes have been implicated in placental adhesion, however, their role remains controversial. This review examines the evidence for and against the involvement of var genes in placental adhesion, and considers whether the most appropriate vaccine candidates have yet been identified.
Association of upper and lower gastrointestinal tract symptoms with body mass index in an Australian cohort
Authors: Talley N. J. , Quan C. , Jones M. P. , Horowitz M.
Published: August 2004

Abstract  Food modulates gastrointestinal (GI) function and GI symptoms could alter food intake, but it is not established whether or not obese people experience more or less GI symptoms. We aimed at evaluating the association between body mass index (BMI) and specific GI symptoms in the community. Population‐based random samples from Sydney, Australia (n = 777) completed a validated questionnaire. The association of each GI symptom with BMI (kg m−2) categories was assessed using logistic regression analysis adjusting for potential confounders. The prevalence of obesity (BMI ≥30 kg m−2) was 22%. There were univariate associations (adjusting for age, sex, education level, alcohol and smoking) between increased BMI category and heartburn (OR = 1.9, 95% CI 1.4, 2.5), acid regurgitation (OR = 2.1, 95% CI 1.4, 2.9), increased bloating (OR = 1.3, 95%CI 1.1, 1.6), increased stool frequency (OR = 1.4, 95% CI 1.1, 1.7), loose and watery stools (OR = 1.5, 95% CI 1.1, 2.0) and upper abdominal pain (OR = 1.3, 95% CI 1.03, 1.6). Early satiety was associated with a lower BMI category but this was not significant after adjustment (OR = 0.8, 95% CI 0.6, 1.1). Lower abdominal pain, postprandial fullness, nausea and vomiting were not associated with BMI category. In a regression model adjusting for sex, education, smoking, alcohol and all GI symptoms, older age, less early satiety and increased stool frequency and heartburn were all independently associated with increasing BMI (all P < 0.01). Heartburn and diarrhoea were associated with increased BMI, while early satiety was associated with a lower BMI in this population.
Global changes in transcription orchestrate metabolic differentiation during symbiotic nitrogen fixation in Lotus japonicus
Authors: Colebatch Gillian , Desbrosses Guilhem , Ott Thomas , Krusell Lene , Montanari Ombretta , Kloska Sebastian , Kopka Joachim , Udvardi Michael K.
Published: July 2004

Research on legume nodule metabolism has contributed greatly to our knowledge of primary carbon and nitrogen metabolism in plants in general, and in symbiotic nitrogen fixation in particular. However, most previous studies focused on one or a few genes/enzymes involved in selected metabolic pathways in many different legume species. We utilized the tools of transcriptomics and metabolomics to obtain an unprecedented overview of the metabolic differentiation that results from nodule development in the model legume, Lotus japonicus. Using an array of more than 5000 nodule cDNA clones, representing 2500 different genes, we identified approximately 860 genes that were more highly expressed in nodules than in roots. One‐third of these are involved in metabolism and transport, and over 100 encode proteins that are likely to be involved in signalling, or regulation of gene expression at the transcriptional or post‐transcriptional level. Several metabolic pathways appeared to be co‐ordinately upregulated in nodules, including glycolysis, CO2 fixation, amino acid biosynthesis, and purine, haem, and redox metabolism. Insight into the physiological conditions that prevail within nodules was obtained from specific sets of induced genes. In addition to the expected signs of hypoxia, numerous indications were obtained that nodule cells also experience P‐limitation and osmotic stress. Several potential regulators of these stress responses were identified. Metabolite profiling by gas chromatography coupled to mass spectrometry revealed a distinct metabolic phenotype for nodules that reflected the global changes in metabolism inferred from transcriptome analysis.
Metabolic engineering of ketocarotenoid formation in higher plants
Authors: Ralley Louise , Enfissi Eugenia M.A. , Misawa Norihiko , Schuch Wolfgang , Bramley Peter M. , Fraser Paul D.
Published: July 2004

Although higher plants synthesize carotenoids, they do not possess the ability to form ketocarotenoids. In order to generate higher plants capable of synthesizing combinations of ketolated and hydroxylated carotenoids the genes responsible for the carotene 4,4′ oxygenase and 3,3′ hydroxylase have been transformed into tomato and tobacco. The gene products were produced as a polyprotein. Subsequent cleavage of the polyprotein, targeting of the two enzymes to the plastid and enzyme activities have been shown for both gene products. Metabolite profiling has shown the formation of ketolated carotenoids from β‐carotene and its hydroxylated intermediates in tobacco and tomato leaf. In the nectary tissues of tobacco flowers a quantitative increase (10‐fold) as well as compositional changes were evident, including the presence of astaxanthin, canthaxanthin and 4‐ketozeaxanthin. Interestingly, in this tissue the newly formed carotenoids resided predominantly as esters. These data are discussed in terms of metabolic engineering of carotenoids and their sequestration in higher plant tissues.
Isolation and identification of phosphatidic acid targets from plants
Authors: Testerink Christa , Dekker Henk L. , Lim Ze‐Yi , Johns Melloney K. , Holmes Andrew B. , de Koster Chris G. , Ktistakis Nicholas T. , Munnik Teun
Published: July 2004

Phosphatidic acid (PA) is emerging as an important lipid signalling molecule. In plants, it is implicated in various stress‐signalling pathways and is formed in response to wounding, osmotic stress, cold stress, pathogen elicitors, Nod factors, ethylene and abscisic acid. How PA exerts its effects is still unknown, mainly because of the lack of characterized PA targets. In an approach to isolate such targets we have used PA‐affinity chromatography. Several PA‐binding proteins were present in the soluble fraction of tomato andArabidopsiscells. Using mass spectrometric analysis, several of these proteins, including Hsp90, 14‐3‐3 proteins, an SnRK2 serine/threonine protein kinase and the PP2A regulatory subunit RCN1 could be identified. As an example, the binding of one major PA‐binding protein, phosphoenolpyruvate carboxylase (PEPC), was characterized further. Competition experiments with different phospholipids confirmed specificity for PA. Hypo‐osmotic treatment of the cells increased the amount of PEPC that bound the PA beads without increasing the absolute amount of PEPC. This suggests that PEPC's affinity for PA had increased. The work shows that PA‐affinity chromatography/mass spectrometry is an effective way to isolate and identify PA‐binding proteins from plants.
Functional interactions between a glutamine synthetase promoter and MYB proteins
Authors: Gómez‐Maldonado Josefa , Avila Concepción , de la Torre Fernando , Cañas Rafael , Cánovas Francisco M. , Campbell Malcolm M.
Published: July 2004

In Scots pine (Pinus sylvestris), ammonium assimilation is catalysed by glutamine synthetase (GS) [EC 6.3.1.2], which is encoded by two genes, PsGS1a and PsGS1b. PsGS1b is expressed in the vascular tissue throughout the plant body, where it is believed to play a role in recycling ammonium released by various facets of metabolism. The mechanisms that may underpin the transcriptional regulation of PsGS1b were explored. The PsGS1b promoter contains a region that is enriched in previously characterized cis‐acting elements, known as AC elements. Pine nuclear proteins bound these AC element‐rich regions in a tissue‐specific manner. As previous experiments had shown that R2R3‐MYB transcription factors could interact with AC elements, the capacity of the AC elements in the PsGS1b promoter to interact with MYB proteins was examined. Two MYB proteins from loblolly pine (Pinus taeda), PtMYB1 and PtMYB4, bound to the PsGS1b promoter were able to activate transcription from this promoter in yeast, arabidopsis and pine cells. Immunolocalization experiments revealed that the two MYB proteins were most abundant in cells previously shown to accumulate PsGS1b transcripts. Immunoprecipitation analysis and supershift electrophoretic mobility shift assays implicated these same two proteins in the formation of complexes between pine nuclear extracts and the PsGS1b promoter. Given that these MYB proteins were previously shown to have the capacity to activate gene expression related to lignin biosynthesis, we hypothesize that they may function to co‐regulate lignification, a process that places significant demands on nitrogen recycling, and GS, the major enzyme involved in the nitrogen recycling pathway.
The S haplotype‐specific F‐box protein gene, SFB, is defective in self‐compatible haplotypes of Prunus avium and P. mume
Authors: Ushijima Koichiro , Yamane Hisayo , Watari Akiko , Kakehi Eiko , Ikeda Kazuo , Hauck Nathanael R. , Iezzoni Amy F. , Tao Ryutaro
Published: July 2004

Many Prunus species, including sweet cherry and Japanese apricot, of the Rosaceae, display an S‐RNase‐based gametophytic self‐incompatibility (GSI). The specificity of this outcrossing mechanism is determined by a minimum of two genes that are located in a multigene complex, termed the S locus, which controls the pistil and pollen specificities. SFB, a gene located in the S locus region, encodes an F‐box protein that has appropriate S haplotype‐specific variation to be the pollen determinant in the self‐incompatibility reaction. This study characterizes SFBs of two self‐compatible (SC) haplotypes, S4′ and Sf, of Prunus. S4′ of sweet cherry is a pollen‐part mutant (PPM) that was produced by X‐ray irradiation, while Sf of Japanese apricot is a naturally occurring SC haplotype that is considered to be a PPM. DNA sequence analysis revealed defects in both SFB4′ and SFBf. A 4 bp deletion upstream from the HVa coding region of SFB4′ causes a frame‐shift that produces transcripts of a defective SFB lacking the two hypervariable regions, HVa and HVb. Similarly, the presence of a 6.8 kbp insertion in the middle of the SFBf coding region leads to transcripts for a defective SFB lacking the C‐terminal half that contains HVa and HVb. As all reported SFBs of functional S haplotypes encode intact SFB, the fact that the partial loss‐of‐function mutations in SFB are present in SC mutant haplotypes of Prunus provides additional evidence that SFB is the pollen S gene in GSI in Prunus.
From pollen tubes to infection threads: recruitment of Medicago floral pectic genes for symbiosis
Authors: Rodríguez‐Llorente Ignacio D. , Pérez‐Hormaeche Javier , Mounadi Kaoutar El , Dary Mohammed , Caviedes Miguel A. , Cosson Viviane , Kondorosi Adam , Ratet Pascal , Palomares Antonio J.
Published: July 2004

While the biology of nitrogen‐fixing root nodules has been extensively studied, little is known about the evolutionary events that predisposed legume plants to form symbiosis with rhizobia. We have studied the presence and the expression of two pectic gene families in Medicago, polygalacturonases (PGs) and pectin methyl esterases (PMEs) during the early steps of the Sinorhizobium meliloti–Medicago interaction and compared them with related pollen‐specific genes. First, we have compared the expression of MsPG3, a PG gene specifically expressed during the symbiotic interaction, with the expression of MsPG11, a highly homologous pollen‐specific gene, using promoter‐gus fusions in transgenic M. truncatula and tobacco plants. These results demonstrated that the symbiotic promoter functions as a pollen‐specific promoter in the non‐legume host. Second, we have identified the presence of a gene family of at least eight differentially expressed PMEs in Medicago. One subfamily is represented by one symbiotic gene (MtPER) and two pollen‐expressed genes (MtPEF1 and MtPEF2) that are clustered in the M. truncatula genome. The promoter‐gus studies presented in this work and the homology between plant PGs, together with the analysis of the PME locus structure and MtPER expression studies, suggest that the symbiotic MsPG3 and MtPER could have as ancestors pollen‐expressed genes involved in polar tip growth processes during pollen tube elongation. Moreover, they could have been recruited after gene duplication in the symbiotic interaction to facilitate polar tip growth during infection thread formation.
Transcriptional profiling by cDNA‐AFLP and microarray analysis reveals novel insights into the early response to ethylene in Arabidopsis
Authors: De Paepe Annelies , Vuylsteke Marnik , Van Hummelen Paul , Zabeau Marc , Van Der Straeten Dominique
Published: July 2004

A comprehensive transcriptome analysis by means of cDNA‐amplified fragment length polymorphism (AFLP) and cDNA‐microarray technology was performed in order to gain further understanding of the molecular mechanisms of immediate transcriptional response to ethylene. Col‐0 plants were treated with exogenous ethylene and sampled at six different time‐points ranging from 10 min until 6 h. In order to isolate truly ethylene‐responsive genes, both the ethylene‐insensitive mutant ein2‐1 and the constitutive mutant (ctr1‐1) were analysed in parallel by cDNA‐AFLP while ein2‐1 was included for the microarray experiment. Out of the cDNA‐transcript profiling covering about 5% of the Arabidopsis transcriptome, 46 ethylene‐responsive genes were isolated, falling in different classes of expression pattern and including a number of novel genes. Out of the 6008 genes present on the chip, 214 genes were significantly (α = 0.001) differentially expressed between Col‐0 and ein2‐1 over time. Cluster analysis and functional grouping of co‐regulated genes allowed to determine the major ethylene‐regulated classes of genes. In particular, a large number of genes involved in cell rescue, disease and defence mechanisms were identified as early ethylene‐regulated genes. Furthermore, the data provide insight into the role of protein degradation in ethylene signalling and ethylene‐regulated transcription and protein fate. Novel interactions between ethylene response and responses to several other signals have been identified by this study. Of particular interest is the overlap between ethylene response and responses to abscisic acid, sugar and auxin. In conclusion, the data provide unique insight into early regulatory steps of ethylene response.
Kinetics of labelling of organic and amino acids in potato tubers by gas chromatography‐mass spectrometry following incubation in 13C labelled isotopes
Authors: Roessner‐Tunali Ute , Liu JunLi , Leisse Andrea , Balbo Ilse , Perez‐Melis Alicia , Willmitzer Lothar , Fernie Alisdair R.
Published: July 2004

Metabolic pathways of primary metabolism of discs isolated from potato tubers were evaluated by the use of a gas chromatography‐mass spectrometry (GC‐MS) method generated specifically for this purpose. After testing several possible methods including chemical ionization, it was decided for reasons of sensitivity, reproducibility and speed to use electron impact ionization‐based GC‐MS analysis. The specific labelling and label accumulation of over 30 metabolites including a broad number of sugars, organic and amino acids was analysed following the incubation of tuber discs in [U‐13C]glucose. The reproducibility of this method was similar to that found for other GC‐MS‐based analyses and comparison of flux estimates from this method with those obtained from parallel, yet less comprehensive, radiolabel experiments revealed close agreement. Therefore, the novel method allows quantitatively evaluation of a broad range of metabolic pathways without the need for laborious (and potentially inaccurate), chemical fractionation procedures commonly used in the estimation of fluxes following incubation in radiolabelled substrates. As a first experiment the GC‐MS method has been applied to compare the metabolism of wild type and wel‐characterized transgenic potato tubers exhibiting an enhanced sucrose mobilization. The fact that this method is able to rapidly yield further comprehensive information into primary metabolism illustrates its power as a further phenotyping tool for the analysis of plant metabolism.
Antisense downregulation of the barley limit dextrinase inhibitor modulates starch granule size distribution, starch composition and amylopectin structure
Authors: Stahl Yvonne , Coates Steve , Bryce James H. , Morris Peter C.
Published: July 2004

The barley protein limit dextrinase inhibitor (LDI), structurally related to the α‐amylase/trypsin inhibitor family, is an inhibitor of the starch debranching enzyme limit dextrinase (LD). In order to investigate the function of LDI, and the consequences for starch metabolism of reduced LDI activity, transgenic barley plants designed to downregulate LDI by antisense were generated. Homozygous antisense lines with reduced LDI protein level and activity were analysed and found to have enhanced free LD activity in both developing and germinating grains. In addition the antisense lines showed unpredicted pleiotropic effects on numerous enzyme activities, for example, α‐ and β‐amylases and starch synthases. Analysis of the starch showed much reduced numbers of the small B‐type starch granules, as well as reduced amylose relative to amylopectin levels and reduced total starch. The chain length distribution of the amylopectin was modified with less of the longer chains (>25 units) and enhanced number of medium chains (10–15 units). These results suggest an important role for LDI and LD during starch synthesis as well as during starch breakdown.