bluuebunny/crossref_metadata_2025_split huggingface dataset:
Potential mechanisms regulating the extracellular activities of basic fibroblast growth factor (FGF‐2)
Authors: Baird Andrew 
Published: February 2005

AbstractBasic fibroblast growth factor (FGF‐2) is nearly ubiquitous in its distribution, leading numerous investigators to propose that there must exist highly specific mechanisms to regulate its bioavailability. Moreover, in each of the tissues where it can be localized, numerous cells are its potential target. The identification of these mechanisms could serve as an important step in developing novel strategies to inhibit FGF action. It could be possible to block such FGF‐dependent activities as angiogenesis, tumor growth, reproduction, and selected diseases of cell proliferation. Over the course of the past several years, we have attempted to describe some of the processes that might regulate a target cell's ability to activate FGF‐2 in its local milieu. © 1994 Wiley‐Liss, Inc.
Structure‐function studies of FGF‐1: Dissociation and partial reconstitution of certain of its biological activities
Authors: Burgess Wilson H. , Friesel Robert , Winkles Jeffrey A. 
Published: February 2005

AbstractWe reported previously that the mitogenic activities of FGF‐1 (acidic FGF) could be dissociated from its receptor‐binding activities by site‐directed mutagenesis of lysine 132 to a glutamic acid. Although the mutant FGF‐1 protein binds to the high‐affinity tyrosinekinase receptors, stimulates tyrosine‐kinase activity, and promotes expression of immediate‐early genes, it is not mitogenic for a variety of tested cell lines. Interestingly, the mutant FGF‐1 is capable of other functions associated with the wild‐type protein such as promotion of mesoderm formation in Xenopus animal caps. The mutant exhibits a reduced apparent affinity for heparin‐Sepharose compared to the wild‐type protein. The relationship between the reduced heparin affinity and lack of mitogenic activity of this mutant is not clear. Recent data indicates the relationship is not as simple as reduced stability of the protein. When NIH 3T3 cells are transfected with expression vectors encoding either wild‐type or mutant FGF‐1, a transformed phenotype can be seen in cells overexpressing the wild‐type FGF‐1, whereas cells overexpressing mutant FGF‐1 appear normal. Analysis of lysates of these cells indicates that a tyrosine‐kinase cascade, distinct from that associated with the high‐affinity cell surface receptors, has been activated in the wild‐type transfected cells but not in the mutant transfected cells. Although both transfected cell lines contain FGF‐1 cell surface receptors as judged by crosslinking studies, the wild‐type transfects are refractory to exogenous FGF‐1, whereas the mutant transfectants respond normally. Together these results support an intracellular role of wild‐type FGF‐1 in mediating certain of its functions. In addition, they demonstrate that certain functions of the growth factor can be dissociated at the structural level. Additional mutagenesis studies have resulted in the identification of mutants with heparin‐binding or mitogenic deficiencies that do not correlate as well as those of the 132 mutant. It appears that the inactivity of the lysine 132 mutant is related, in part, to cysteine 131. © 1994 Wiley‐Liss, Inc.
Heparan sulfate fibroblast growth factor receptor complex: Structure‐function relationships
Authors: McKeehan Wallace L. , Kan Mikio 
Published: February 2005

AbstractSplice variations in genes coding for the transmembrane FGF receptor (FGFR) result in isoforms that vary in the ectodomain, intracellular juxtamembrane domain, and the intracellular kinase domain. An analysis of biochemical functions of distinct recombinant isoforms expressed in baculoviral‐infected insect cells allowed generation of models for function of splice variants in both the ecto‐ and intracellular domains. A structural model for the ectodomain of the FGFR is proposed as follows. Alternately‐spliced immunoglobulin‐like disulfide Loop I, which is not required for ligand‐binding, is sufficiently interactive with the base FGF binding site formed by Loops II and III to modify ligand affinity and affect interaction of the receptor with heparan sulfate cofactor. The NH2‐terminal domain of Loop II, which is highly conserved across all isoforms, exhibits a 19‐residue heparin‐binding domain which is obligatory for FGF binding. Heparin protects a 30‐kDa ligand‐binding fragment from proteolysis that is composed of Loop II, the inter‐Loop II/III sequence, and the NH2‐terminus of Loop III. This suggests that the high‐affinity FGF receptor complex is an intimate ternary complex of transmembrane tyrosine kinase, heparan sulfate glycosaminoglycan, and FGF, each of which have interactive binding domains for the other and may contribute to specificity of the FGFR complex. Although Ig Loop II, the inter‐Loop II/III sequence, and the NH2‐terminus of Loop III with heparan sulfate form the base FGF binding site, mutually exclusive alternate splicing of two exons coding for the COOH‐terminal half of Loop III determines which specific members of the FGF ligand family bind with high affinity to the base site.A kinase‐ and tyrosine phosphorylation site‐defective splice variant, FGFR type 2, acts as a dominant‐negative suppressor of phosphorylation of specifically tyr‐653 in the catalytic domain of the kinase, with less effect on phosphorylation of tyr‐766 in the COOH‐terminal tail. We propose that phosphorylation of tyr‐766, which is required for interaction of phospholipase Cγ1 (PLCγ1) with the receptor, may occur by a cis‐intramolecular mechanism within FGFR monomers, while phosphorylation of tyr‐653, which is required for phosphorylation of PLCγ1, may occur by a trans‐intermolecular mechanism between monomers within kinase homodimers. From the combined results, we propose a model whereby increasing concentrations of FGF may control FGF‐mediated signal transduction by heterodimerization of different FGFR monomers. Different monomers arise by regulated combinatorial alternate splicing that alters both the extracellular and intracellular domains. © 1994 Wiley‐Liss, Inc.
Onset of paternal gene activation in early mouse embryos fertilized with transgenic mouse sperm
Authors: Matsumoto Kazuya , Anzai Masayuki , Nakagata Naomi , Takahashi Akio , Takahashi Yumi , Miyata Kenji 
Published: February 2005

AbstractWe investigated the onset of paternal gene expression in the early mouse embryo. We obtained transgenic mouse embryos by fertilizing BD (C57BL/6N x DBA) F1 hybrid female oocytes in vitro, with sperm from homozygous transgenic males carrying integrated chicken β‐actin promoter‐driven firefly luciferase cDNA. We then examined the RNA and protein synthesis of the luciferase gene in embryos from the 1‐ to 2‐cell stage. RNA transcripts of the luciferase gene were first detected in the 1‐cell stage embryos as early as 13 hr postinsemination, just prior to elongation. By photon‐count imaging, functional luciferase was identified at the 2‐cell stage 23 hr postinsemination. These findings indicate that the paternal endogenous gene is already transcribed in the late 1‐cell embryos, although paternally derived protein is not synthesized until the 2‐cell stage. Therefore, these results suggest that the embryonic gene is activated as early as the late 1‐cell stage. ©1994 Wiley‐Liss, Inc.
Transfer of nuclei from 8‐cell stage mouse embryos following use of nocodazole to control the cell cycle
Authors: J. Otaegui P. , T. O'Neill G. , Campbell K. H. S. , Wilmut I. 
Published: February 2005

AbstractMouse 2‐, 4‐, 8‐, and 16‐cell embryos were exposed to nocodazole in M16 culture medium. The effect of different concentrations and exposure times on the efficiency of cell cycle synchronization and the development of the treated embyros after release from the drug was determined. The minimum effective concentration (95% of arrested nuclei) for 4‐, 8‐, and 16‐cell embryos was 5μM nocodazole. The effect upon subsequent development of mouse embryos depended upon both the stage of development of the embryo at treatment (P < 0.001) and the length of exposure to nocodazole (P < 0.001). Exposure to any concentration of nocodazole within the range 2.5–10 μM for 12 hr caused a reduction in the proportion of embryos that formed blastocysts. As the period of exposure to 5μM nocodazole increased from 12 to 24 hr, the proportion of embryos developing to the blastocyst stage decreased. The lower proportion of embyros developing to the blastocyst stage and to term (P < 0.01) suggests that the more advanced stages were more susceptible to damage as a result of exposure to nocodazole. The rate of development of 4‐cell embryos to blastocysts was not affected when an exposure time of 9 hr was used. Together these results show that it is possible to use nocodazole to arrest mouse embryonic cells in mitosis but that it is not appropriate to culture the embryos in the presence of this drug for prolonged periods. Individual blastomeres completed mitosis at 60–90 min and started DNA synthesis at 120–150 min after release from nocodazole. Nuclei from blastomeres thus synchronized were used to conduct studies on the effect of the cell cycle on nuclear transfer. A signficant effect was found. When nuclei from 8‐cell embryos in G1 or S‐phase were used as nuclei donors, development to blastocyst was respectively 27% and none. ©Wiley‐Liss, Inc.
Formation of the vitelline envelope precedes the active uptake of vitellogenin during oocyte development in the rainbow trout,Oncorhynchus mykiss
Authors: Johan Hyllner Sven , Silvers Christer , Haux Carl 
Published: February 2005

AbstractIn order to study the initial formation of the vitelline envelope and the appearance of vitellogenin in oocytes of rainbow trout, females were sampled monthly from 19 to 5 mo before ovulation. lmmunohistochemistry revealed that the formation of the vitelline envelope starts when the oocytes reach a diameter of about 450 μm. Oocytes of this size were first found in females sampled a year before ovulation at the time when plasma levels of estradiol‐17β increased from 0.2 to 0.6 ng/ml. An antiserum directed against vitellogenin crossreacted with small vesicles (around 2 μm) present just inside the oolemma, when the oocytes reached a diameter of 600 μm. This was interpreted as an active uptake of vitellogenin. Oocytes of this size were first found in females sampled 9 mo before ovulation at the time when estradiol 17β levels increased from 0.6 to 1.0 ng/ml and the gonadal somatic index was doubled. Oocytes with a diameter of 600 μm had an immunoreactive vitelline envelope with a thickness of about 3 μm. It is apparent that the initial formation of the vitelline envelope starts before the active uptake of vitellogenin and that the low previtellogenic plasma levels of estradiol‐17β observed in females are of physiological significance. © 1994 Wiley‐Liss, Inc.
Basal components of the transcription apparatus (RNA polymerase II, TATA ‐binding protein) contain activation domains: Is the repetitive c‐terminal domain (CTD) of RNA polymerase II a “Portable Enhance Domain”?
Authors: Seipel Katja , Georgiev Oleg , Gerber Hans‐Peter , Schhaffner Walter 
Published: February 2005

AbstractRegions rich in serine, threonine, and proline residues can be found in transcriptional activation domains, as well as in the N‐terminal parts of mammalian TATA‐binding proteins, where they are interrupted by polyglutamine stretches. Likewise, the C‐terminal domain of the largest subunit of RNA polymerase II contains multiple repeats of the consensus heptapeptide sequence YSPTSPS. To test directly for possible activation functions, we fused the GAL4 DNA‐binding domain to the N‐terminal domain of human TBP or subdomains of it, and to the C‐terminal domain (CTD) of mouse RNA polymerase II or synthetic polymers of a CTD consensus repeat. We found that these chimeric proteins were able to activate transcription when bound to a GAL4 site in front of the TATA box, a function characteristic of transcription factors. However, while subdomains of TBP functioned only from a position close to the TATA box (“promoter” position), multiple repeats of the CTD consensus sequence were also able to mediate transcriptional activation from a remote (“enhancer”) position.Our findings suggest that a region of TBP that is unique to mammals functionally cooperates with “proximal” activation domains of promoter‐bound transcription factors. They also imply that the C‐terminal domain of RNA polymerase II includes a function that is otherwise confined to remote activation domains of enhancer‐bound transcription factors. We suggest that the CTD of RNA polymerase II contains a “portable” remote activation domain that may also facilitate chromatin opening within the transcription unit. © 1994 Wiley‐Liss, Inc.
Transforming growth factor‐β1 in reproduction
Authors: M. Shull Marcia , Doetschman Thomas 
Published: February 2005

AbstractExpression patterns of TGF‐βs during embryogenesis and in adult reproductive organs, as well as the activities of these molecules in in vitro assays of biological processes relating to reproduction and development, have suggested that TGF‐βs may play a role in both reproductive function and embryonic development. To investigate the function of TGF‐β1 in vivo, the murine TGF‐β1 gene was disrupted by gene targeting, and animals that lacked TGF‐β1 activity were generated. Homozygous mutant animals were obtained which exhibited a multifocal inflammatory disease. However, the observed numbers of homozygous mutant offspring were less than expected, suggesting the occurrence of some type of prenatal lethality. This paper reviews the proposed role of the TGF‐βs in reproductive and developmental processes and discusses observations obtained from the TGF‐β1 gene‐targeting experiments as they relate to these processes. © 1994 Wiley‐Liss, Inc.
Analysis of thymidine kinase gene expression in preimplantation mouse embryos
Authors: Lee Dae Kee , Sun Woong , Cho Hyeseong , Kim Kyungjin , Rhee Kunsoo , Lee Chung Choo 
Published: February 2005

AbstractThymidine kinase (TK) activity was examined during the development of preimplantation mouse embryos. TK activity was increased approximately 20‐fold from day 2 embryos (2‐cell) to day 5 embryos (late blastocyst). TK activity did not change along with the progression into S‐phase of the first and the second cell cycles but increased sharply at S‐phase of the third cell cycle. Analysis of TK mRNA with a reverse transcription‐polymerase chain reaction (RT‐PCR) method showed that the level of TK mRNA was low in ovulated eggs and 1‐cell embryos and was hardly detectable in day 2 embryo (2‐cell), but sharply increased in day 3 embryos (mixture of 5‐ to 8‐cell and morula). The functional role of 5′‐flanking sequence of TK gene was also investigated in preimplantation embryos after microinjection with the DNA construct of 5′‐flanking sequence of TK (2.4 kb) linked to bacterial lacZ gene (TK2.5lacZ) into the pronucleus of 1‐cell and subsequently by histochemical staining with X‐gal. β‐Galactosidase activity was first detected in day 3 embryos (8‐cell), and 30% of embryos were stained with X‐gal in day 4 and day 5 embryos, respectively. These results show that an increase in TK activity occurred after 2‐cell stage, and this increase was primarily due to the embryonic activation of TK gene expression. Also, it appears that the 5′‐flanking sequence of TK may directly regulate the TK gene expression at the transcriptional level during preimplantation murine development. © 1994 Wiley‐Liss, Inc.
Immunoglobulins from human follicular fluid induce the acrosome reaction in human sperm
Authors: Saragüeta Patricia , Lanuza Guillermo , V. Miranda Patricia , G. Tezón Jorge , Lino Baraño J. 
Published: February 2005

AbstractThe ability of human follicular fluid (hFF), retrieved from women undergoing IVF to induce the acrosome reaction (AR) in human sperm has been documented by several laboratories. However, the nature of the active factors in the hFF and the physiological meaning of the AR induction are highly controversial. We performed a three step purification scheme for hFF and all the fractions were screened for the AR‐inducing activity. AR activity was associated with a protein fraction of Mr > 180 kD that on further analysis under PAGE was found to be composed by subunits of apparent Mr 50,000 and 29,000. The N‐terminal sequences of these bands showed a 100% homology with the heavy and light chains of human lgG. A polyclonal antibody raised against the purified protein and anti‐human lgG were both able to suppress the acrosome reactioninducing activity of crude hFF. However, neither normal human serum nor a purified preparation of human lgG were able to mimic the AR‐inducing activity of hFF. We concluded that the AR‐inducing activity of hFF is, at least in part, due to the presence of antisperm antibodies. © 1994 Wiley‐Liss, Inc.
Calcium influx into mouse spermatozoa activated by solubilized mouse zona pellucida, monitored with the calcium fluorescent indicator, fluo‐3. inhibition of the influx by three inhibitors of the zona pellucida induced acrosome reaction: Tyrphostin A48, pertussis toxin, and 3‐quinuclidinyl benzilate
Authors: L. Bailey Janice , T. Storey Bayard 
Published: February 2005

AbstractThe fluorescent calcium indicator, fluo‐3, was loaded as the membrane permeant tetraacetoxymethyl (AM) ester into cauda epididymal mouse sperm at 25°C for 20 min in the absence of bovine serum albumin (BSA) and presence of the dispersant, Pluronic F‐127. Excess indicator was removed by two centrifugation washes at 100g for 10 min, a procedure that did not impair sperm motility. Upon resuspension in medium containing 20 mg/ml BSA to promote capacitation, the sperm cells exhibited readily detectable fluorescence uniformly distributed in the cytoplasm. Cell fluorescence was stable over the time of the experiments and was responsive to changes in intracellular calcium concentration, [Ca2+]j. Initial [Ca2+]j was 231 ± 58 nM (±SE, n = 43). Addition of heat‐solubilized mouse zonae pellucidae to capacitated sperm increased [Ca2+]j by 106 ± 19 nM (±SE, n = 18), the higher steady‐state concentration being reached after 30 min. Subsequent addition of the non‐fluorescent calcium ionophore Br‐A23187 resulted in a further increase of 114 ± 18 nM (± SE, n = 18), the higher steady‐state concentration being reached after 6 min. The increase in [Ca2+]j induced by solubilized zonae pellucidae was largely blocked by 3‐quinuclidinyl benzilate (QNB) an antagonist of muscarinic receptors that was earlier shown to block the zona pellucida induced acrosome reaction in mouse sperm (Florman and Storey, 1982: Dev Biol 91:121–130). This [Ca2+]j increase was completely blocked by the tyrosine kinase inhibitor, tyrphostin A48, and by the inactivator of G1 proteins, pertussis toxin. At the concentrations at which they blocked the zona pellucida‐induced increase in [Ca2+]j all three inhibitors also blocked the zona pellucidainduced acrosome reaction. These results indicate that [Ca2+]j increase in is an early, if not the initial, reaction in the sequence leading to zona pellucida induced acrosomal exocytosis in mouse sperm. The observation that the three inhibitors, each having a different mode of action, all block the zona pellucida induced [Ca2+]j suggests that the sperm plasma membrane receptors mediating the zona pellucida induced acrosome reaction may function as a complex, whose formation is activated by zona pellucida ligand binding. © 1994 Wiley‐Liss, Inc.
Developmental expression of poly(A) binding protein mRNAs during spermatogenesis in the mouse
Authors: C. Kleene K. , Wang M‐Y. , Cutler M. , Hall C. , Shih D. 
Published: February 2005

AbstractThe poly(A) binding protein (PABP), a conserved protein that binds to the 3′ poly(A) tail on mRNAs in eukaryotic cells, has been implicated in the regulation of mRNA stability and translation. Two PABP cDNAs with different sequences were isolated from mouse testis cDNA libraries. The predicted amino acid sequence of one, PABP1, is nearly identical (98.9%) to human liver PABP, while 80% of the amino acids of the second, PABPt, are identical to mouse and human PABPs. Northern blots reveal that there is one major PABP mRNA species in liver, muscle, kidney, and brain, two in spleen, and at least four in testis. The levels of PABP mRNA in testis are 5–10‐fold higher than in these somatic tissues, but surprisingly the vast majority of all PABP mRNA size variants sediment more slowly than single ribosome, indicating strong translational repression. Reverse transcriptase‐polymerase chain reaction assays demonstrate that PABPt mRNAs are abundant only in testis. Northern blots of RNAs purified from highly enriched spermatogenic cells show that the high levels, multiple sizes of PABP mRNAs, and the PABPt mRNA are present in meiotic and early haploid spermatogenic cells, and are sharply reduced in late haploid cells. Comparison of the binding of PABP1 and PABPt to poly(A) Sepharose in vitro revealed subtle differences, even though PABPt contains substitutions for highly conserved aromatic amino acids that are thought to be necessary for binding to poly(A). The existence of two PABP isoforms in mouse spermatogenic cells could influence cytoplasmic gene expression during spermatogenesis. © 1994 Wiley‐Liss, Inc.
Timetable of in vivo embryonic development in the grey short‐tailed opossum (Monodelphis domestica)
Authors: E. Mate Karen , Robinson Edward S. , A. Pedersen Roger , L. Vandeberg John 
Published: February 2005

AbstractThe timing of development was examined in 496 embryos from female Monodelphis domestica, collected at known time intervals after video recorded mating. Ovulation occurred approximately 20 hr (day 1) after mating, and fertilization was observed by 24 hr. Transport through the oviducts was rapid, and pronuclear stage embryos were recovered from the uterus as early as 24 hr after mating. Second cleavage had occurred by 55 hr after mating. Three‐celled embryos were among those collected on day 3 after mating, indicating that asynchronous cleavage of blastomeres can occur from the two‐cell stage. The four‐cell stage persisted for approximately 24 hr, and embryos that had undergone third cleavage were first recovered 74 hr after mating. Embryos that had undergone fourth to fifth cleavage were found 96–100 hr (4 days) after mating and complete unilaminar blastocysts by 5.5 days after mating. Primary endoderm formed from an already distinct embryonic area of the unilaminar blastocyst early on day 7 after mating. Formation of the bilaminar blastocyst was completed rapidly, on day 7 after mating. The primitive streak appeared on day 10 after mating, and organogenesis rapidly ensued on a timetable similar to that reported for Didelphis virginiana (McCrady, 1938). Close contact with the maternal circulation was established on day 11 and by day 12 maternal and embryonic tissues could not be separated without damage. The length of the gestation period from fertilization to birth was approximately 13.5 days. These observations provide the basis for further embryological cellular and molecular studies of this species as a laboratory model for marsupial development.© 1994 Wiley‐Liss, Inc.