Biomedical subjects

Michael D Sheets

Publications and source records attributed to Michael D Sheets.

9 recordsLinked to original sources

PubMed2007

Chordin affects pronephros development in Xenopus embryos by anteriorizing presomitic mesoderm.

Spemann's organizer emits signals that pattern the mesodermal germ layer during Xenopus embryogenesis. In a previous study, we demonstrated that FGFR1 activity within the organizer is required for the production of both the somitic muscle- and pronephros-patterning signals by the organizer and the expression of chordin, an organizer-specific secreted protein (Mitchell and Sheets [2001] Dev. Biol. 237:295-305). Studies from others in both chicken and Xenopus embryos provide compelling evidence that pronephros forms by means of secondary induction signals emitted from anterior somites (Seufert et al. [1999] Dev. Biol. 215:233-242; Mauch et al. [2000] Dev. Biol. 220:62-75). Here we provide several lines of evidence in support of the hypothesis that chordin influences pronephros development by directing the formation of anterior somites. Chordin mRNA was absent in ultraviolet (UV) -irradiated embryos lacking pronepheros (average DAI<2) but was always found in UV-irradiated embryos that retain pronepheros (average DAI>2). Furthermore, ectopic expression of chordin in embryos and in tissue explants leads to the formation of anterior somites and pronephros. In these experiments, pronephros was only observed in association with muscle. Chordin diverted somatic muscle cells to more anterior positions within the somite file in chordin-induced secondary trunks and induced the expression of the anterior myogenic gene myf5. Finally, depletion of chordin mRNA with DEED antisense oligonucleotides substantially reduced somitic muscle and pronephric tubule and duct formation in whole embryos. These data and previous studies on ectoderm and endoderm (Sasai et al. [1995] Nature 377:757) support the idea that chordin functions as an anteriorizing signal in patterning the germ layers during vertebrate embryogenesis. Our data support the hypothesis that chordin directs the formation of anterior somites that in turn are necessary for pronephros development.

Animals↗

PubMed2006

Heading in a new direction: implications of the revised fate map for understanding Xenopus laevis development.

Amphibian embryos have served as a model system for vertebrate axial patterning for more than a century. Recent changes to the Xenopus laevis fate map revised the assignment of the embryonic dorsal/ventral (back-to-belly) axis in pre-gastrula embryos and allowed the assignment of the rostral/caudal (head-to-tail) axis for the first time. Revising the embryonic axes after many years of experimentation changes our view of axial patterning in amphibians. In this review, we discuss the revised maps and axes, and show by example how the new map alters the interpretation of three experiments that form the foundations of amphibian embryology. We compare the revised amphibian fate map to the general maps of the protochordates, and discuss which features of the maps and early development are shared by chordates and which distinguish vertebrates. Finally, we offer an explanation for the formation of both complete and incomplete axes in the rescue assays routinely used to study axial patterning in Xenopus, and a model of amphibian axial patterning.

Animals↗

PubMed2005

Fate mapping hematopoietic lineages in the Xenopus embryo.

Fate maps reveal the normal contribution of various regions of an early embryo to larval and adult structures. This chapter describes the mapping of primitive blood in embryos of the frog Xenopus laevis by the technique of lineage tracing with a fluorescent dextran.

Animals↗

PubMed2004

BMP antagonism by Spemann's organizer regulates rostral-caudal fate of mesoderm.

Recent revisions to the Xenopus fate map challenge the interpretation of previous maps and current models of amphibian axial patterning (Lane, M.C., Smith, W.C., 1999. The origins of primitive blood in Xenopus: implications for axial patterning. Development 126 (3), 423-434.; Lane, M.C., Sheets, M.D., 2000. Designation of the anterior/posterior axis in pregastrula Xenopus laevis. Dev. Biol. 225, 37-58). We determined the rostralmost contributions to both dorsal and ventral mesoderm concomitantly from marginal zone progenitors in stage 6 embryos. Data reveal an unequivocal rostral-to-caudal progression of both dorsal and ventral mesoderm across the pre-gastrula axis historically called the dorsal-ventral axis, and a dorsal-to-ventral progression from animal-to-vegetal in the marginal zone. These findings support the proposed revisions to the fate and axis orientation maps. Most importantly, these results raise questions about the role of the organizer grafts and organizer-derived BMP antagonists in the "induction" of secondary axes. We re-examine both phenomena, and find that organizer grafts and BMP antagonists evoke caudal-to-rostral mesodermal fate transformations, and not ventral-to-dorsal transformations as currently believed. We demonstrate that BMP antagonism evokes a second axis because it stimulates precocious mediolateral intercalation of caudal, dorsal mesoderm. The implications of these findings for models of organizer function in vertebrate axial patterning are discussed.

Animals↗

PubMed2004

Xenopus embryonic poly(A) binding protein 2 (ePABP2) defines a new family of cytoplasmic poly(A) binding proteins expressed during the early stages of vertebrate development.

We describe a new RNA binding protein from Xenopus we have named ePABP2 (embryonic poly(A) binding protein type II). Based on amino acid similarity, ePABP2 is closely related to the ubiquitously expressed nuclear PABP2 protein that directs the elongation of mRNA poly(A) tails during pre-mRNA processing. However, in contrast to known PABP2 proteins, Xenopus ePABP2 is a cytoplasmic protein that is predominantly expressed during the early stages of Xenopus development and in adult ovarian tissue. Biochemical experiments indicate ePABP2 binds poly(A) with specificity and that this binding requires the RRM domain. Mouse and human ePABP2 proteins were also identified and mouse ePABP2 expression is also confined to the earliest stages of mouse development and adult ovarian tissue. We propose that Xenopus ePABP2 is the founding member of a new class of poly(A) binding proteins expressed in vertebrate embryos. Possible roles for this protein in regulating mRNA function in early vertebrate development are discussed.

Aging↗

PubMed2003

Expression of scFv antibodies in Xenopus embryos to disrupt protein function: implications for large-scale evaluation of the embryonic proteome.

SUMMARY: We evaluated the use of single-chain antibody (scFv) expression as a tool to disrupt the function of specific proteins in embryos of the frog, Xenopus laevis. The expression of scFvs that recognize the bone morphogenetic protein receptor (ALK3) or the fibroblast growth factor receptor1 (FGFR1) as endoplasmic reticulum-anchored proteins caused distinct developmental defects that were virtually indistinguishable from the defects caused by expression of the dominant negative forms of each receptor. These results demonstrate that scFvs from phage-display libraries can be readily fashioned into effective and specific inhibitors of signaling pathways in developing embryos. In addition, as several effective scFvs against a specific target can be isolated rapidly, this approach represents a valuable new tool for large-scale functional analysis of the embryonic proteome.

Activin Receptors, Type I↗

PubMed2002

Primitive and definitive blood share a common origin in Xenopus: a comparison of lineage techniques used to construct fate maps.

Primitive blood constitutes the ventralmost mesoderm in amphibians, and its cleavage-stage origin reveals important clues about the orientation of the dorsal/ventral axis in the embryo. In recent years, investigators employing various lineage-labeling strategies have reported disparate results for the origin of primitive blood in Xenopus [W. D. Tracey, Jr., M. E. Pepling, G. H. Thomsen, and J. P. Gergen (1998). Development 125, 1371-1380; M. C. Lane W. C. Smith (1999). Development 126, 423-434; K. R. Mills, D. Kruep, and M. S. Saha (1999). Dev. Biol. 209, 352-368; A. Ciau-Uitz, M. Walmsley, and R. Patient (2000). Cell 102, 787-796]. These discrepancies must be resolved in order to elucidate early embryonic patterning mechanisms in vivo. We directly compared two of the techniques used to determine the origin of the ventral blood islands and primitive blood, injection of either beta-galactosidase mRNA or conjugated dextrans, by coinjecting both tracers simultaneously into individual blastomeres in cleavage-stage embryos. We find that dextrans label progeny efficiently, while beta-galactosidase activity is not present in many of the progeny of an injected blastomere, suggesting that mRNA fails to diffuse throughout a blastomere. This result demonstrates that beta-galactosidase mRNA fails to meet the criterion for a true lineage label, namely efficient detection of the progeny of a blastomere, and raises questions about interpretations based on mapping the ventral blood islands using Lac Z mRNA as a tracer. We examined the origins of the ventral blood islands and primitive blood from the vegetal region of the marginal zone in regularly cleaving embryos by coinjecting both reporters into C-tier blastomeres. Our results demonstrate that both the ventral blood islands and primitive blood routinely arise from all C-tier blastomeres. Our data, in combination with published mapping results for the dorsal aorta, demonstrate that primitive and definitive blood do not have separate origins at the 32-cell stage in Xenopus. In addition, these results support a proposal to align the dorsal/ventral axis of the mesendoderm with the animal/vegetal axis in pregastrula Xenopus.

Animals↗

PubMed2002

Rethinking axial patterning in amphibians.

Recent revisions in the Xenopus laevis fate map led to the designation of the rostral/caudal axis and reassignment of the dorsal/ventral axis (Lane and Smith [1999] Development 126:423-434; Lane and Sheets [2000] Dev. Biol. 225:37-58). It is unprecedented to reassign primary embryonic axes after many years of research in a model system. In this review, we use insights about vertebrate development from anatomy and comparative embryology, as well as knowledge about gastrulation in frogs, to reexamine several traditional amphibian fate maps. We show that four extant maps contain information on the missing rostral/caudal axis. These maps support the revised map as well as the designation of the rostral/caudal axis and reassignment of the dorsal/ventral axes. To illustrate why it is important for researchers to use the revised map and nomenclature when thinking about frog and fish embryos, we present an example of alternative interpretations of "dorsalized" zebrafish mutations.

Amphibians↗

PubMed2002

Zygotic control of maternal cyclin A1 translation and mRNA stability.

Cyclin mRNAs are unstable in the adult cell cycle yet are stable during the first 12 cell divisions in Xenopus laevis. We recently reported that cyclin A1 and B2 maternal mRNAs are deadenylated upon completion of the 12th division (Audic et al. [2001] Mol. Cell Biol. 21:1662-1671). Deadenylation is mediated by the 3' untranslated region (UTR) of the mRNA and precedes the terminal disappearance of the cyclin proteins, with both processes requiring zygotic transcription. The purpose of the current study was (1) to ask whether deadenylation leads to translational repression and/or destabilization of endogenous cyclin A1 and B2 mRNAs, and (2) to further characterize the regulatory sequences required. We show that zygote-driven deadenylation leads to translational repression and mRNA destabilization. A 99-nucleotide region of the 3'UTR of the cyclin A1 mRNA mediates both deadenylation and destabilization. Surprisingly, two AU-rich consensus elements within this region are dispensable for this activity. These results suggest that zygote-dependent deadenylation, translational repression, and mRNA destabilization by means of novel 3'UTR elements contribute to the disappearance of maternal cyclins. They also suggest that translational control of cyclins may play a role in the transition to the adult cell cycle. These data concur with previous studies in Drosophila showing that zygote-mediated degradation of maternal cdc25 mRNA may be a general mechanism whereby transition to the adult cell cycle proceeds.

3' Untranslated Regions↗