bluuebunny/crossref_metadata_2025 huggingface dataset:
Genetics of sex determination in flowering plants
Authors: Grant Sarah , Houben Andreas , Vyskot Boris , Siroky Jiri , Pan Wei‐Hua , Macas Jiri , Saedler Heinz
Published: May 2005

Most flowering plant species are hermaphroditic, but a small number of species in most plant families are unisexual (i.e., an individ‐ual will produce only male or female gametes). Because species with unisexual flowers have evolved repeatedly from hermaphroditic progenitors, the mechanisms controlling sex determination in flowering plants are extremely diverse. Sex is most strongly determined by genotype in all species but the mechanisms range from a single controlling locus to sex chromosomes bearing several linked locirequired for sex determination. Plant hormones also influence sex expression with variable effects from species to species. Here, we review the genetic control of sex determination from a number of plant species to illustrate the variety of extant mechanisms. We emphasize species that are now used as models to investigate the molecular biology of sex determination. We also present our own investigations of the structure of plant sex chromosomes of white campion (Silene latifolia ‐ Melan‐drium album). The cytogenetic basis of sex determination in white campion is similar to mammals in that it has a male‐specific Y‐chromosome that carries dominant male determining genes. If one copy of this chromosome is in the genome, the plant is male. Otherwise it is female. Like mammalian Y‐chromosomes, the white campion Y‐chromosome is rich in repetitive DNA. We isolated repetitive sequences from microdissected Y‐chromosomes of white campion to study the distribution of homologous repeated sequences on the Y‐chromosome and the other chromosomes. We found the Y to be especially rich in repetitive sequences that were generally dispersed over all the white campion chromosomes. Despite its repetitive character, the Y‐chromosome is mainly euchromatic. This may be due to the relatively recent evolution of the white campion sex chromosomes compared to the sex chromosomes of animals. © 1994 Wiley‐Liss, Inc.
Function of the her‐1 gene is required for maintenance of male differentiation in adult tissues of C. elegans
Authors: Schedin Pepper , Jonas Petra , Wood William B.
Published: May 2005

Function of the sex‐determining gene her‐1 is required in XO embryos cf C. elegans to specify male development. Using a temperature‐sensitive mutant of her‐1, we show that when XO males reared at a permissive temperature are shifted as adults to a nonpermissive temperature, they initiate vitellogenin synthesis in the intestine and oocyte production in the germline. A similar shift has no effect on her‐1(+) males. We conclude that sexual differentiation of the intestine and germline is plastic, requiring her‐1 expression throughout adulthood for maintenance of the male state. © 1994 Wiley‐Liss, Inc.
Behavioral and neurobiological implications of sex‐determining factors inDrosophila
Authors: Taylor Barbara J. , Villella Adriana , Ryner Lisa C. , Baker Bruce S. , Hall Jeffrey C.
Published: May 2005

The function of the central nervous system as it controls sex‐specific behaviors inDrosophilahas been studied with renewed intensity, in the context of genetic factors that influence the development of sexually differentiated aspects of this insect. Three categories of genetic variations that cause anomalies in courtship and mating behaviors are discussed: (1) mutants isolated with regard to courtship defects, of which putatively courtship‐specific variants such as thefruitlessmutant are a subset; (2) general behavioral and neurological variants (including sensory and learning mutants), whose defects include subnormal reproductive performance; and (3) mutations of genes within the sex‐determination regulatory hierarchy ofDrosophila, the analysis of which has included studies of reproductive behavior. Recent studies of mutations in two of these categories have provided new insights into the control of neuronally based aspects of sex‐specific behavior. Thedoublesexgene, the final factor acting in the sex‐determination hierarchy, had been previously thought to regulate all aspects of sexual differentiation. Yet, it has been recently shown thatdoublesexdoes not control at least one neuronally‐determined feature of sex‐specific anatomy—a muscle in the male's abdomen, whose normal development is, however, dependent on the action offruitless.These considerations prompted us to examine further (and in some cases re‐examine) the influences exerted by sex‐determination hierarchy genes on behavior. Our results—notably those obtained from assessments ofdoublesexmutations' effects on general reproductive actions and on a particular component of the courtship sequence (male “singing” behavior)—lead to the suggestion that there is a previously unrecognized branch within the sexdetermination hierarchy, which controls the differentiation of the male‐ and female‐ specific phenotypes ofDrosophila.This new branch separates from thedoublesex‐related one immediately before the action of that gene (just afterfransformerandtransformer‐2) and appears to control as least some aspects of neuronally determined sexual differentiation of males. © 1994 Wiley‐Liss, Inc.
Temperature‐dependent sex determination in reptiles: Proximate mechanisms, ultimate outcomes, and practical applications
Authors: Crews David , Bergeron Judith M. , Bull James J. , Flores Deborah , Tousignant Alan , Skipper James K. , Wibbels Thane
Published: May 2005

In many egg‐laying reptiles, the incubation temperature of the egg determines the sex of the offspring, a process known as temperature‐dependent sex determination (TSD). In TSD sex determination is an “all or none” process and intersexes are rarely formed. How is the external signal of temperature transduced into a genetic signal that determines gonadal sex and channels sexual development? Studies with the red‐eared slider turtle have focused on the physiological, biochemical, and molecular cascades initiated by the temperature signal. Both male and female development are active processes—rather than the crganized/default system characteristic of vertebrates with genotypic sex determination—that require simultaneous activation and suppression of testis‐ and ovary‐determining cascades for normal sex determination. It appears that temperature accomplishes this end by acting on genes encoaing for steroidogenic enzymes and steroid hormone receptors and modifying the endocrine microenvironment in the embryo. The temperature experienced in development also has long‐term functional outcomes in addition to sex determination. Research with the leopard gecko indicates that incubation temperature as well as steroid hormones serve as organizers in shaping the adult phenotype, with temperature modulating sex hormone action in sexual differentiation. Finally, practical applications of this research have emerged for the conservation and restoration of endangered egg‐laying reptiles as well as the embryonic development of reptiles as biomarkers to monitor the estrogenic effects of common environmental contaminants. © 1994 Wiley‐Liss, Inc.
S6 phosphorylation results from prothoracicotropic hormone stimulation of insect prothoracic glands: A role for S6 kinase
Authors: Song Qisheng , Gilbert Lawrence I.
Published: May 2005

The insect prothoracic glands are the source of steroidal molting hormone precursors and the glands are stimulated by a brain neuropeptide, prothoracicotropic hormone (PTTH). Previous work from this laboratory revealed that PTTH acts via a cascade including Ca2+/calmodulin activation of adenylate cyclase, protein kinase A, and the subsequent phosphorylation of a 34 kDa protein (p34) hypothesized, but not proven, to be the 56 protein of the 40S ribosomal subunit. The jmmunosuppressive macrolide, rapamycin, is a potent inhibitor of cell proliferation, a signal transduction blocker, and also prevents ribosomal S6 phosphorylation in mammalian systems. We demonstrate here that rapamycin inhibited PTTH‐stimulated ecdysteroidogenesis in vitro by the prothoracic glands of the tobacco hornworm, Manduca sexta, with half‐maximal inhibition at a concentration of about 5 nM. At concentrations above 5 nM, there was a 75% inhibition of ecdysteroid biosynthesis. Similar results, were observed with the calcium ionophore (A23187), a known stimulator of ecdysteroidogenesis. Most importantly, the inhibition of ecdysteroid biosynthesis was accompanied by the specific inhibition of the phosphorylation of p34, indicating that p34 indeed is ribosomal protein S6. In vivo assays revealed that injection of rapamycin into day 6 fifth instar larvae resulted in a decreased hemolymph ecdysteroid titer and a dose‐dependent delay in molting and metamor‐phosis. When S6 kinase (S6K) activity was examined using rapamycin‐treated prothoracic glands as the enzyme source and a synthetic peptide (S6‐21) or a 40S ribosomal subunit fraction from Manduca tissues as substrate, the date revealed that rapamycin inhibited S6K activity. The composite data suggest that rapamycin inhibits a signal transduction element leading to p34 phosphorylation that is necessary for PTTH‐stimulated ecdysteroidogenesis in this insect endocrine gland, and lend further support to the concept that p34 is S6. © 1994 Wiley‐Liss, Inc.
Cloning of a thyroid hormone‐responsive Rana catesbeiana c‐erbA‐β gene
Authors: Davey Jennifer C. , Schneider Mark J. , Galton Valerie Anne
Published: May 2005

Two types of thyroid hormone receptor (c‐erbA) gene have been identified in mammals and in lower species including chickens and the amphibian Xenopus laevis. The two genes are located on different chromosomes and have been named TRα and TRβ We have described previously the cloning of a TRβ cDNA from Rana catesbeiana (RC) tissues (RC15) and we now report the cloning of a TRβ cDNA from this species. The cloning strategy employed utilized the polymerase chain reaction (PCR), with primers based on the sequences of the X. laevis TRβ cDNA (XenTRβ) and an RCTRβ genomic clone, which, by analogy with XenTRβ, contains some of the 3′ end of the open reading frame together with 3′‐untranslated sequences. At the nucleotide and amino acid levels, respectively, the cloned RCTRβ cDNA is 90% and 98% homologous with XenTRβ, and 72% and 76% homologous with RC15. Following in vitro transcription and translation, the cDNA was shown to encode a 48 kilodalton protein which binds 3,5, 3′‐triiodothyronine (T3) with high affinity (mean Kd: 0.032 nM). Samples of total or poly(A)+RNA from tadpoles at different stages of metamorphosis and from adult frogs were analyzed for the presence ofTRβ‐specific transcripts by slot blot analysis using as probe a 258 bp section of the RCTRβ cDNA. This section of the cDNA does not hybridize to the corresponding section of RC15. In confirmation of previous findings, β‐specific transcripts were not detected in RNA from tadpole red blood cells (RBCs) and none was found in RBCs from adult frogs. However, β‐specific transcripts were detected in RNA from tail, skin, and liver of stage XIII tadpoles, and the levels were greatly increased in these tissues during metamorphic climax. In the adult frog, the level was minimal in all tissues studied. In other studies using stage XII tadpoles and a reverse transcription‐polymerase chain reaction assay, β‐specific transcripts were readily detected in tail, skin, kidney, leg, heart, intestine, and eye but minimal or absent in liver, brain, and RBCs. Within 2 days of injection of T3, the level of β‐specific transcripts was markedly increased in all tissues. These findings strongly suggest that the expression of the RCTRβ gene is regulated developmentally and/or by thyroid hormone in most tissues of the RC tadpole. © 1994 Wiley‐Liss, Inc.
Upregulation of AP‐2 in the skin of Xenopus laevis during thyroid hormone‐induced metamorphosis
Authors: French Randall P. , Warshawsky David , Tybor Lisa , Mylniczenko Natalie D. , Miller Leo
Published: May 2005

During amphibian metamorphosis dramatic changes occur in the morphogenesis and differentiation of the epidermis. Concurrently with these changes, the 63 kDa keratin geneis upregulated from low basal levels to high levels. What makes these processes unique is that they are controlled by triiodothyronine (T3) and can be duplicated in cultures of purified epidermal cells. Since there is a 2 day lag period between the addition of T3 and the upregulation of keratin gene expression and terminal differentiation, recent studies have focused on identifying the genes activated during the lag period. We assume that the transcription factors required for upregulation of the keratin gene are induced by T3 during the lag period, and therefore we have cloned the keratin gene so that promoter analyses can be conducted. S1 mapping assays have shown that the same transcription start sites are used during premetamorphosis when the keratin gene is basally expressed, during metamorphosis when it is T3‐upregulated, and in the adult epidermis where it is expressed independently of T3. During the early part of the lag period TRP and AP‐2 mRNA levels are upregulated in the epidermis by T3. The transcription factor AP‐2 is expressed at high levels in the skin of premetamorphic larvae and induced about fivefold by T3 but is not induced in an epithelial cell line (XL‐177). Since the keratin mRNA, AP‐2 rnRNA, and other genes induced during the lag period are expressed in premetamorphic larvae it appears that T3 functions by upregulating the expression of genes previously activated by a T3‐independent process. This preprogramming may account for the tissue specificity of T3 action during metamorphosis. © 1994 WiIey. Liss, Inc.
frizzled Gene expression and development of tissue polarity in the Drosophila wing
Authors: Park Woo Jin , Liu Jingchun , Adler Paul N.
Published: May 2005

Almost every cell in the Drosophila pupal wing forms a single, distally pointing cuticular hair. The function of the frizzled (fz) gene is essential for the elaboration of the normal wing hair pattern. In the absence of fz function hairs develop, but they display an abnormal polarity. We have examined the developmental expression of the fi gene at the RNA level via in situ hybridization and at the protein level via Western blotting. We have found that fz is expressed in all regions of the epidermis before, during, and after the fz cold sensitive period. We have also found that fz function is not required for normal fi expression. We have further found that mutations in several other tissue polarity genes do not noticeably alter the expression or the modification state of the Fz protein. © 1994 Wiley‐Liss, Inc.
TheKnottedleaf blade is a mosaic of blade, sheath, and auricle identities
Authors: Sinha Neelima , Hake Sarah
Published: May 2005

The dominantKnotted‐1mutations in maize alter development of the leaf blade. Sporadic patches of localized growth, or knots, and fringes of ectopic ligule occur along lateral veins of mutant leaf blades. In addition, bundle sheaths do not completely encircle lateral veins on mutant leaf blades. We have compared mutant leaf blades with wild‐type leaves to determine the precise nature of the perturbed regions. Our analysis includes characterization of epidermal cell shapes, localization of photosynthetic proteins and histology of the leaf. We show that mutant leaf blades are a mosaic of leaf organ components. Affected regions of mutant leaf blades resemble either sheath or auricle tissue in both external and internal features. This conversion of blade cells represents an acropetal shift of more basal parts of the leaf blade region and correlates with previously identified ectopic expression of theKnotted‐1protein in the leaf blade. We propose that inappropriate expression ofKn1interferes with the process of establishment of cell identities, resulting in early termination of the normal blade development program or precocious expression of the sheath and auricle development programs. © 1994 Wiley‐Liss, Inc.
Epigenetic effects in eukaryotic gene expression
Authors: Bestor Timothy H. , Chandler Vicki L. , Feinberg Andrew P.
Published: May 2005

In the broadest terms, epigenetic phenomena in eukaryotes depend on the interaction of alleles or repeated sequences or on the mitotic inheritance of chromatin states or methylation patterns. One of the most exciting aspects of the study of epigenetic phenomena is the insight that can be gained into the structure and assembly of higher‐order chromatin structures, an important subject that has proved refractory to current biochemical methodologies. Rapid progress in the study of gene inactivation in fungi, plants, and invertebrates will provide new hypotheses to be tested in mammals. © 1994 Wiley‐Liss, Inc.
Spreading the silence: Epigenetic transcriptional regulation during Drosophila development
Authors: Moehrle Axel , Paro Renato
Published: May 2005

In early Drosophila development a complex cascade of diffusible transcription factors generates an intricate expression pattern of developmental regulators such as the homeotic genes. The mechanism which subsequently maintains the pattern during the rest of development is mainly using epigenetic features for its function. Evidence comes from the analysis of the Polycomb‐group (Pc‐G), a class of genes which is responsible for maintaining the inactive state of expression. The Pc‐G was found to share many parallels to genes involved in heterochromatin formation. Different members of the Pc‐G interact in large multiprotein complexes, which apparently can cover and inactivate large chromosomal domains. Specific DNA elements have been identified that are used by the Pc‐G proteins to nucleate these specialized domains of silent chromatin. Thus, the Pc‐G proteins appear to permanently inactivate genes by generating heterochromatin‐like structures which could then be inherited by the daughter cells in an epigenetic manner. Heritable gene silencing is an important but little understood mechanism in pattern formation. Phenomenologically related effects have been observed in many organisms. These range from the transcriptional silencing of the inactive mating type loci in yeast to parental imprinting phenomena and X‐chromosome inactivation in mammals. Analysis of these functions in Drosophila provides an excellent model system for studying the molecular basis of such epigenetic mechanisms that use higher order chromatin structures for transcriptional repression. © 1994 Wiley‐Liss, Inc.
Positional information and pattern formation in development
Authors: Wolpert L.
Published: May 2005

A widely used mechanism for pattern formation is based on positional information: cells acquire positional identities as in a coordinate system and then interpret this information according to their genetic constitution and developmental history. In Drosophila maternal factors establish the axes and set up a maternal system of positional information on which further patterning is built. There is a cascade of gene activity which leads both to the development of periodic structures, the segments, and to their acquiring a unique identity. This involves the binding of transcription factors to regulatory regions of genes to produce sharp thresholds. Many of the genes involved in these processes, particularly the Hox complex, are also involved in specifying the body axis and limbs of vertebrates. There are striking similarities in the mechanisms for spcifying and recording positional identity in Drosophila and vertebrates. © 1994 Wiley‐Liss, Inc.
Developmental significance of epigenetic impositions on the plant genome: A paragenetic function for chromosomes
Authors: Jorgensen Richard
Published: May 2005

Developmental and physiological factors can impose heritable metastable changes on the plant genome, a fact that was established by maize geneticists during the 1950s and 1960s, largely through the efforts of R. Alexander Brink and Barbara McClintock. This paper describes a transgenic reporter system that monitors genomic impositions as changes in morphogenetically‐determined flower color patterns. The observations reported here on the metastable properties of plant transgenes illustrate the proposals of Brink and McClintock that chromosomal impositions occur during normal development as ordered sequences of events which contribute to the elaboration of complex developmental patterns. The relationship between this process and some recent findings about the control of gene expression in transgenic plants is also discussed. © 1994 Wiley‐Liss, Inc.
Tumor suppression in Drosophila is causally related to the function of the lethal(2)tumorous imaginal discs gene, a dnaJ homolog
Authors: Kurzik‐Dumke Ursula , Gundacker Dietmar , Rentrop Martin , Gateff Elisabeth
Published: May 2005

The Drosophila melanogaster tumor suppressor gene lethal(2)tumorous imaginal discs (l(2)tid) causes in homozygotes malignant growth of cells of the imaginal discs and the death of the mutant larvae at the time of puparium formation. We describe the molecular cloning of the 1(2)tid+ gene and its temporal expression pattern in the wild‐type and mutant alleles. Germ line rescue of the tumor phenotype was achieved with a 7.0 kb Hindlll‐fragment derived from the polytene chromosome band 59F5. The l(2)tid+ gene spans approximately 2.5 kb of genomic DNA. The protein coding region, 1,696 bps long, is divided by an intron into two exons. The predicted Tid56 protein contains 518 amino acids and possesses a theoretical molecular weight of 56 kDa. It shows significant homology to all known DnaJ related proteins from bacteria, yeast, and man. The possible function of the Tid56 protein in tumor suppression is delineated. © 1995 Wiley‐Liss, Inc.
Epidermal cell‐specific quantitation of dopa decarboxylase mRNA in Drosophila by competitive RT‐PCR: An effect of Broad‐Complex mutants
Authors: O'Keefe Sandra , Schouls Michelle , Hodgetts Ross
Published: May 2005

The quontitation of RNA in tissue homogenates by amplifying the product of reverse transcription (RT‐PCR) is sufficiently sensitive to detect molecules in the range of 10−110−2 amole. We describe here the steps we believe necessary to validate a protocal that used a DNA competitor and visualization of the amplification products by ethidium bromide staining. The procedure was designed to quantitate one of the tissue specific transcripts of the Dopa decarboxylase gene (Ddc) in Drosophila. We demonstrate that the amount of epidermal Ddc transcript is much lower at pupariation in several mutants of the Broad‐Complex, one of the primary response loci of the moulting hormone, ecdysone. The mutant effects were allele specific and the molecular basis of one of these alleles is known. This implicates a particular family of the zinc finger proteins encoded by the locus in the hormone dependent induction of Ddc expression.
Germ cell‐sertoli cell interactions: Regulation by germ cells of the stage‐specific expression of CP‐2/cathepsin LmRNA by sertoli cells
Authors: Wright William W. , Zabludoff Sonya D. , Penttilä Tarja‐Leena , Parvinen Martti
Published: May 2005

CP‐2/cathepsin LmRNA is expressed primarily by rat Sertoli cells within stage VI–VIII seminiferous tubules. To test whether germ cells regulated this expression, we examined if separating Sertoli cells from specific germ cells affected expression of this transcript in Sertoli cells. First, Sertoli cells were isolated from adult (90‐day‐old) and immature (25‐day‐old) rats and levels of this transcript measured immediately or after 1, 3 and 5 days in culture. Results demonstrated that immediately upon isolation, CP‐2/cathepsin LmRNA levels were significantly higher in mature cells. However, after 1 day in culture, the levels of this transcript increased in immature cells and remained high in mature cells. We therefore conclude that in vivo, a subset of germ cells inhibit the expression of CP‐2/cathepsin LmRNA by immature Sertoli cells. Second, to examine the effect of specific germ cells on CP‐2/cathepsin LmRNA expression, we exposed the testes of mature rats to 3 Gy of γ‐radiation and analyzed stage‐specific expression of this transcript at varying times during maturation depletion and subsequent germ cell restoration. Loss of spermatogonia or spermatocytes was without effect. However, when pachytene spermatocytes through step 14 spermatids were depleted, expression at stages VI–VIII was reduced by half and expression at stages IX‐I was increased 14‐fold. These changes resulted in the loss of stage‐specific expression of CP‐2/cathepsin LmRNA by Sertoli cells. Finally, stage VI–VIII tubules, depleted primarily in step 15–19 spermatids, had levels of CP‐2/cathepsin LmRNA that were 60% of control. However, stage‐specific expression of this transcriot was detected in these tubules. In contrast to whai we noted with CP‐2/cathepsin LmRNA, loss and restoration of germ cells had no effect on Sertoli cell levels of SGP‐2 mRNA, indicating that testicular irradiation had no Overall effect on Sertoli cell function Taken togetherr these data suggest that the stage‐specific expression of the CP‐2/ cathepsin L gene results from the sequential stimulation and inhibition of Sertoli cells by germ cells, that pachytene spermatocytes through step 14 spermatids are required for this stage‐specific expression and that step 18 and 19 spermatids amplify this expression at stages VI–VIII. © 1995 Wiley‐Liss, Inc.
Transcriptional regulation of Sertoli cell differentiation (transferrin promoter activation) during testicular development
Authors: Chaudhary Jaideep , Skinner Michael K.
Published: May 2005

Previously testicular peritubular cells have been shown to produce a paracrine factor PModS that promotes Sertoli cell differentiation. This mesenchymal‐epithelial cell interaction appears to regulate a number of Sertoli cell differentiated functions including transferrin gene expression. The current study was designed to identify PModS‐activated response elements in the transferrin promoter and correlate this with Sertoli cell differentiation that occurs during testis development. The 3‐kb transferrin promoter was digested down to approximately 200‐bp fragments. Nuclear extracts from Sertoli cells stimulated with PModS were used in gel mobility shift assays. Two promoter regions located at −2.4 kb and −1.9 kb were designated SE1 and SE2. PModS promoted the presence of factors in Sertoli cell nuclear extracts that bind SE1 and SE2. Displacement studies demonstrated that SE1 and SE2 are distinct. A transferrin promoter‐reporter construct containing these apparent response elements was activated by PModS, while a minimal transferrin promoter of 600bp excluding SE1 and SE2 was only partially stimulated by PModS. Therefore, PModS appears to in part activate the transferrin promoter through SE1 and/or SE2. Gel shift assays with Sertoli cell nuclear extracts and 20‐day‐old testis extracts were the same. Interestingly, the nuclear extract from a new‐born testis also had a gel shift. Therefore, some of the nuclear factors stimulated by PModS in Sertoli cells and present in mid‐pubertal testis were also present at birth upon completion of embryonic development. Previously transferrin expression has been shown to increase significantly at the onset of puberty. Observations indicate that PModS appears to in part promote transferrin expression through two newly identified response elements designated SE1 and SE2 and that the nuclear factors that bind these elements are present after embryonic development and mid‐pubertally. © 1995 Wiley‐Liss, Inc.
A haploid and a diploid cell cycle coexist in an in vitro immortalized spermatogenic cell line
Authors: Hofmann Marie‐Claude , Abramian Donara , Millán José Luis
Published: May 2005

We have recently established a conditionally immortalized germ cell line [GC‐2spd(ts)] that, at the permissive temperatures of 37°C and 32°C, is able to undergo meiosis in vitro and form round spermatids [Hofmann et al., (1994): Proc Natl Acad Sci USA 91:5533–5537]. In this report, we provide data that indicate that the GC‐2spd(ts) cell line consists of two cell populations undergoing a haploid (n/2n) cell cycle and a diploid (2n/4n) cell cycle, respectively. The cells containing 2n DNA, when sorted by fluorescence‐activated cell sorting, are able to reconstitute the full population of n/2n/4n DNA cells, indicating that they are able to commit to the reductive meiotic division and form haploid spermatids or to continue self‐replication through a diploid cell cycle. This GC‐2spd(ts) cell line provides a valuable tool to study the molecular mechanisms involved in the cellular decision between self‐renewal by mitosis and commitment to meiosis. © 1995 Wiley‐Liss, Inc.
Distinct patterns of expression of the D‐type cycling during testicular development in the mouse
Authors: Ravnik Stuart E. , Rhee Kunsoo , Wolgemuth Debra J.
Published: May 2005

The three D‐type cyclins have been shown to be differentially expressed in a number of isolated cell types and cell lines, suggesting distinct roles in cell cycle regulation in particular cell lineages. The testis provides unique opportunities to study genes involved in cell cycle regulation, since it contains cells in both mitosis and meiosis as well as differentiated cells with little proliferation activity. Major transcripts of 4.2 kb, 6.8 kb, and 2.3 kb were detected in the adult mouse testis by Northern hybridization analyses for cyclin D1, cyclin D2, and cyclin D3, respectively. Additional transcripts of 1.8 and 2.7 kb were detected by Northern hybridization for cycin D3 in the testis, but not in other tissues, and these transcripts were limited to germ cells. Northern and in situ hybridization analyses of normal and germ cell‐deficient testes showed the surprising result that cyclin D1 was expressed in a pattern consistent with expression in the non‐dividing Sertoli cells. Cyclin D2 levels appeared slightly enriched in germ cell‐deficient testes as compared to intact testis, but in situ hybridization analysis did not reveal any distinct cellular localization. Also surprising was the observation that cyclin D3 expression was highest in the non‐dividing, haploid, round spermatids. The possible roles of these cyclins in the events of spermatogenesis are discussed. © 1995 Wiley‐Liss, Inc.
Genomic organization of a mouse glyceraldehyde 3‐phosphate dehydrogenase gene (Gapd‐s) expressed in post‐meiotic spermatogehic cells
Authors: Welch J. E. , Brown P. R. , O'Brien D. A. , Eddy E. M.
Published: May 2005

The Gapd‐s gene encodes an isoform of the glyceraldehyde 3‐phosphate dehy‐drogenase enzyme expressed only in post‐meiotic spermatogenic cells. Two clones containing the Gapd‐s gene were isolated from a mouse genomic library. Sequencing and restriction enzyme analysis demonstrated that this single‐copy gene contains 11 exons and spans 9596 base pairs. The locations of Gapd‐s exons and introns are conserved when compared to the corresponding portions of the chicken and human somatic Gapd genes. The promoter region contains no TATA box, although there is a potential SP1 recognition site within exon 1. Like other TATA‐less genes, primer extension analysis reveals some heterogeneity in the site of transcription initiation with Gapd‐s transcripts initiating from three discrete sites. Northern analysis demonstrated that a 1.5‐kb Gapd‐s mRNA is expressed in the testis in at least three mammalian orders, indicating that the Gapd‐s gene appeared early in mammalian evolution. Using GAPD‐deficient bacteria, mouse GAPD‐S was shown to be capable of functioning as a glycolytic enzyme. Since GAPD has been proposed to be a key enzyme regulating glycolysis in spermatogenic cells, GAPD‐S may represent a potential target for toxicological or contraceptive agents affecting fertility by interfering with glycolysis. © 1995 Wiley‐Liss, Inc.
The CpG‐rich promoter of human LDH‐C is differentially methylated in expressing and nonexpressing tissues
Authors: Bonny Christophe , Goldberg Erwin
Published: May 2005

A comparison of nucleolide sequences of murine LDH‐a and Ldh‐c genes and human LDH‐A, LDH‐B, and LDH‐C reveals that mouse Ldh‐c has lost the CpG “island” present in the genes for the somatic isozymes. However, the human LDH‐C gene has a CpG‐rich region of 230 bp surrounding its promoter. Endonuclease sensitivity coupled with polymerase chain reaction (PCR) demonstrate the presence of nine heavily methylated sites in this region in different somatic cells. The same sites are specifically hypomethy‐lated in expressing tissues. 3′ sites bordering the CpG‐rich region appear to be methylated in both expressing and nonexpressing tissues. Furthermore, the methylated promoter forms a specific complex in vitro with a methyl‐DNA binding protein. Evolutionary and functional implications of these observations are discussed. © 1995 Wiley‐Liss, Inc.
Developmental regulation of the ovine β‐lactoglobulin/human serum albumin transgene is distinct from that of the β‐lactoglobulin and the endogenous β‐casein genes in the mammary gland of transgenic mice
Authors: Baruch Ariela , Shani Moshe , Hurwitz David R. , Barash Itamar
Published: May 2005

We compared the developmental pattern of expression of the sheep β‐lactoglobulin (BLG), the chimeric BLG/human serum albumin (HSA), and the endogenous murine β‐casein genes in the mammary gland of virgin, pregnant and lactating transgenic mice, both at the RNA (expression) and protein (synthesis and secretion) levels. The BLG and casein genes were expressed at very low levels in virgin animals and during early stages of pregnancy. The increase in the expression of these genes started at the second half of pregnancy and reached a peak between the end of pregnancy and day 10 of lactation. The accumulation of their RNA coincided with that of the corresponding proteins, indicating a transcriptional control of expression of these genes. The expression and secretion patterns of the endogenous casein gene in transgenic and nontransgenic mice were indistinguishable. The hybrid BLG/HSA gene constructs displayed distinct patterns of expression in virgin animals and at early stage of pregnancy, from that of the BLG transgene or the endogenous mouse milk protein gene. High levels of expression (17–60% of that on day 18 of pregnancy) were detected in the mammary gland of virgin animals. At day 5 of pregnancy there was a dramatic decrease in HSA synthesis and secretion in all transgenic strains tested. The down‐regulation, revealed by immunoprecipitation and immunohistochemical studies, demonstrated that at that stage of pregnancy only 10–18% of ductal structures contained HSA expressing cells in contrast to the majority of ducts expressing HSA in virgin animals. These morphological studies also demonstrated that the down‐regulation in HSA synthesis and secretion was correlated with the transition from ducts comprised of a single layer of epithelial cells (characteristic of the virgin state) to ducts composed of multilayers of such cells. In two of the three transgenic strains tested, the down‐regulation at the protein level was associated with a similar decrease in HSA transcripts. In the exceptional strain no. 23, HSA transcripts continued accumulating even at this stage. The differences in the control of expression at the RNA level between these transgenic strains were also confirmed by in situ hybridization. Our results suggest the involvement of at least two regulatory mechanisms effective at early stages of gestation in the control of expression/secretion of the HSA transgene targeted for expression in the mammary gland by the BLG milk protein promoter. These putative mechanisms may play key roles in the interplay between normal mammogenesis and lactogenesis. Thus, transgenic mice expressing BLG/HSA gene constructs at early stages of gestation would be valuable in further dissecting these mechanisms. © 1995 Wiley‐Liss, Inc.
Embryonic regulation of histone ubiquitination the sea urchin
Authors: Jasinskiene Nijole , Jasinskas Algimantas , Langmore John P.
Published: May 2005

We have used quantitative 2‐D protein electrophoresis and immunoprecipitation to study the patterns of histone ubiquitination at 10 h and 36 h of embryonic development in Strongylocentrotus purpuratus. Variants csH2A, αH2A, βH2A, γH2A, δHA, H2AF./Z, αH2B, βH2B, and γH2B showed up to sevenfold differences in level of monoubiquitination between variants, and individual variants showed up to sixfold changes during development. At 36 h of embryogenesis, the late variants were less ubiquitinated than the early variants, althoug h the overall level of ubiquitination was appreciably greater than at 10 h. Antiubiquitin antibodies were used to precipitate formaldehyde‐fixed chromatin fragments in order to estimate the degree of ubiquitination of the early histone genes. The 5′ regulatory region of the active H3 gene appeared to be at least twice as ubiquitinated as the adjacent upstream spacer. However, the absolute level of ubiquitination of the early histone gene repeat seemed to be independent of transcriptional activity. These results show that variant‐specific ubiquitination of histones is a part of the developmental program in sea urchin embryos, but is not clearly correlated with transcriptional activity of the early histone genes, except perhaps in the regulatory regions. © 1995 Wiley‐Liss, Inc.
Phenotypic diversity of 188 rice embryo mutants
Authors: Hong Soon‐Kwan , Aoki Toshiyuki , Kitano Hidemi , Satoh Hikaru , Nagato Yasuo
Published: May 2005

We have identified 188 embryo mutants of rice and characterized them into six groups based on their phenotypes: (1) embryoless in mature seed, (2) deletion of embryonic organ(s), (3) abnormal position of embryonic organs, (4) abnormal embryo size, (5) defect in organ morphology, and (6) variable abnormal phenotypes in spite of single mutations. Three types of organless mutants are obtained: small globular embryo, club‐shaped embryo, and large embryo. Although 12 shootless mutants derived from at least three loci are identified, only three radicleless mutants are recovered, which produce normal adventitious roots after germination. In reduced embryo mutants, every embryonic organ is reduced, in contrast to giant embryo mutants in which only scutellum is enlarged. Considerable number of mutants are categorized into (5) and (6) in the above. These diverse embryo mutants would serve as promising materials for genetic study of embryogenesis. © 1995 Wiley‐Liss, Inc.
Transient reduction in secreted 32 kD chitinase prevents somatic embryogenesis in the carrot (Daucus carota L.) variant ts11
Authors: de Jong Anke J. , Hendriks Theo , Meijer Ellen A. , Penning Maarten , Schiavo Fiorella L. , Terzi Mario , van Kammen Ab , de Vries Sacco C.
Published: May 2005

At the nonpermissive temperature, somatic embryos of the temperature‐sensitive (ts) carrot (Daucus carota L.) cell variant ts11 only proceed beyond the globular embryo stage in the presence of medium conditioned by wild‐type cells. The causative component in the conditioned medium has been identified as an acidic 32 kD endochitinase. An antiserum raised against the 32 kD chitinase detected this protein in culture medium from ts11 embryo cultures grown at the permissive temperature as well as at the nonpermissive temperature. No difference in biochemical characteristics or in effect on ts11 embryo development could be detected between the 32 kD chitinase purified from wild‐type cultures and the chitinase from ts11 cultures grown at the permissive or at the nonpermissive temperature. Compared to the amount present in a ts11 embryo culture at the permissive temperature, a reduction in the amount of 32 kD chitinase was observed during the temperature‐sensitive period at the nonpermissive temperature. These results imply that the arrested embryo phenotype of ts11 is not the result of a structural difference in its 32 kD chitinase, but is the result of a transient decrease in the amount of 32 kD chitinase present. Morphological observations indicate that the ts11 phenotype is pleiotropic and also affects the cell wall of nonembryogenic cells. © 1995 Wiley‐Liss, Inc.
Expression of knotted1 marks shoot meristem formation during maize embryogenesis
Authors: Smith Laurie G. , Jackson David , Hake Sarah
Published: May 2005

The formation of shoot and root meristems that ultimately give rise to all tissues of the plant body occurs for the first time during embryogenesis. Meristem formation has traditionally been defined in terms of the appearance of histological features of meristems; this approach has led to varying interpretations of the timing of meristem formation relative to other events in embryogenesis. Markers that would provide more objective criteria for the analysis of meristem formation have not been widely available. The maize homeobox gene, knotted1 (kn1), is expressed in shoot meristems throughout postembryonic stages of shoot development. In order to determine whether this gene is expressed in the shoot meristem from its earliest inception, we examined the expression of kn1 in embryos at a series of stages by in situ hybridization to kn1 mRNA and immunolocalization of KN1 protein. Our results show that the onset of kn1 expression is temporally and spatially coincident with the earliest histologically recognizable signs of shoot meristem formation in the embryo, and thus provides a valuable marker for this process. © 1995 Wiley‐Liss, Inc.
Regionalisation of cell fate and morphogenetic movement of the mesoderm during mouse gastrulation
Authors: Parameswaran Maala , Tam Patrick P. L.
Published: May 2005

The developmental fate of cells in the epiblast of early‐primitive‐streak‐stage mouse embryos was assessed by studying the pattern of tissue colonisation displayed bylacZ‐expressing cells grafted orthotopically to nontransgenic embryos. Results of these fate‐mapping experiments revealed that the lateral and posterior epiblast contain cells that will give rise predominantly to mesodermal derivatives. The various mesodermal populations are distributed in overlapping domains in the lateral and posterior epiblast, with the embryonic mesoderm such as heart, lateral, and paraxial mesoderm occupying a more distal position than the extraembryonic mesoderm. Heterotopic grafting of presumptive mesodermal cells results in the grafted cells adopting the fate appropriate to the new site, reflecting a plasticity of cell fate determination before ingression. The first wave of epiblast cells that ingress through the primitive streak are those giving rise to extraembryonic mesoderm. Cells that will form the mesoderm of the yolk sac and the amnion make up a major part of the mesodermal layer of the midprimitive‐streak‐stage embryo. Cells that are destined for embryonic mesoderm are still found within the epiblast, but some have been recruited to the distal portion of the mesoderm. By the late‐primitive‐streak‐stage, the mesodermal layer contains only the precursors of embryonic mesoderm. This suggests that there has been a progressive displacement of the midstreak mesoderm to extraembryonic sites, which is reminiscent of that occurring in the overlying endodermal tissue. The regionalisation of cell fate in the late‐primitive‐streak mesoderm bears the same spatial relationship as their ancestors in the epiblast prior to cell ingression. This implies that both the position of the cells in the proximal‐distal axis and their proximity to the primitive streak are major determinants for the patterning of the embryonic mesoderm. © 1995 Wiley‐Liss, Inc.
Mesodermal patterning during avian gastrulation and neurulation: Experimental induction of notochord from non‐notochordal precursor cells
Authors: Yuan Shipeng , Darnell Diana K. , Schoenwolf Gary C.
Published: May 2005

The cells that are normally fated to form notochord occupy a region at the rostral tip of the primitive streak at late gastrula/early neurula stages of avian and mammalian development. If these cells are surgically removed from avian embryos in culture, a notochord will nonetheless form in the majority of cases. The origin of this reconstituted notochord previously had not been investigated and was the objective of this study. Chick embryos at late gastrulal early neurula stages were cultured, and the rostral tip of the primitive streak including Hensen's node was removed and replaced with non‐node cells from quail epiblast to ensure that the cells normally fated to be notochord would be absent and that healing of the blastoderm would occur. Embryos were allowed to develop for 24 hr, and the presence and origin (host or graft) of the notochord were assessed using antibodies against notochord or quail cells. Two notochords typically developed; both were almost exclusively of host origin. The primitive streak, and in some cases adjacent tissues, was removed from another group of embryos in an attempt to estimate the mediolateral position and extent of the cells required to form reconstituted notochord. Additional experimental embryos with and without grafts were transected at various rostrocaudal levels in an attempt to estimate the rostrocaudal extent of the cells required to form reconstituted notochord. Finally, various levels of the primitive streak either were placed in a neutral environment (the germ cell crescent) or were grafted in place of the node. Collective results from all experiments indicate that the areas lateral to the rostral portion of the primitive streak, estimated to have a rostrocaudal span of less than 500 μm and a mediolateral extent of less than 250 μm, are critical for formation of the reconstituted notochord. Fate mapping and histological examination of this region identify 4 possible precursor cell populations. Further studies are underway to determine which of the 4 possible precursor cell types forms or induces the reconstituted notochord, and which tissue interactions underlie this change in cell fate. © 1995 Wiley‐Liss, Inc.
Translational control of activin in Xenopus laevis embryos
Authors: Klein Peter S. , Melton Douglas A.
Published: May 2005

Activin is a potent mesoderm inducing factor present in embryos of Xenopus laevis. Recent evidence has implicated activin in the inhibition of neural development in addition to the well‐established induction of mesoderm in ectodermal explants. These diverse effects are critically dependent on the concentration of activin yet little is known about the mechanisms regulating the level of activin in the embryo. We report that the 3′ untranslated region (3′ UTR) of activin βB mRNA inhibits the translation of activin in embryos. Microinjection of activin mRNA from which the 3′ UTR has been deleted is 8–10‐fold more potent in inducing mesoderm than mRNA containing the 3′ UTR. Truncation of the 3′ UTR also leads to a marked enhancement of activin protein levels in embryos but has no effect when the truncated mRNA is translated in vitro. The 3′ UTR also confers translational inhibition on a heterologous mRNA. These data show that a maternal factor(s) present in X. laevis regulates the translation of injected activin βB mRNA. This factor(s) could be responsible for regulating the levels of endogenous activin βB protein during mesoderm induction and the specification of ectodermal derivatives such as neural and epidermal tissues. © 1995 Wiley‐Liss, Inc.