In JoVE (1)

Other Publications (5)

Articles by Tripti Gupta in JoVE

Other articles by Tripti Gupta on PubMed

Cct1, a Phosphatidylcholine Biosynthesis Enzyme, is Required for Drosophila Oogenesis and Ovarian Morphogenesis

Development (Cambridge, England). Dec, 2003  |  Pubmed ID: 14597574

Patterning of the Drosophila egg requires cooperation between the germline cells and surrounding somatic follicle cells. In order to identify genes involved in follicle cell patterning, we analyzed enhancer trap lines expressed in specific subsets of follicle cells. Through this analysis, we have identified tandem Drosophila genes homologous to CTP: phosphocholine cytidylyltransferase (CCT), the second of three enzymes in the CDP-choline pathway, which is used to synthesize phosphatidylcholine. Drosophila Cct1 is expressed at high levels in three specific subsets of follicle cells, and this expression is regulated, at least in part, by the TGF-beta and Egfr signaling pathways. Mutations in Cct1 result in a number of defects, including a loss of germline stem cell maintenance, mispositioning of the oocyte, and a shortened operculum, suggesting that Cct1 plays multiple roles during oogenesis. In addition, Cct1 mutants display a novel branched ovariole phenotype, demonstrating a requirement for this gene during ovarian morphogenesis. These data provide the first evidence for a specific role for CCT, and thus for phosphatidylcholine, in patterning during development.

Tbx2b is Required for Ultraviolet Photoreceptor Cell Specification During Zebrafish Retinal Development

Proceedings of the National Academy of Sciences of the United States of America. Feb, 2009  |  Pubmed ID: 19179291

The vertebrate rod and cone photoreceptors are highly specialized sensory neurons that transduce light into the chemical and electrical signals of the nervous system. Although the physiological properties of cones and rods are well known, only a handful of genes have been identified that regulate the specification of photoreceptor subtypes. Taking advantage of the mosaic organization of photoreceptors in zebrafish, we report the isolation of a mutation resulting in a unique change in photoreceptor cell fate. Mutation of the lots-of-rods (lor) locus results in a near one-for-one transformation of UV-cone precursors into rods. The transformed cells exhibit morphological characteristics and a gene-expression pattern typical of rods, but differentiate in a temporal and spatial pattern consistent with UV-cone development. In mutant larvae and adults, the highly ordered photoreceptor mosaic is maintained and degeneration is not observed, suggesting that lor functions after the specification of the other photoreceptor subtypes. In genetic chimeras, lor functions cell-autonomously in the specification of photoreceptor cell fate. Linkage analysis and genetic-complementation testing indicate that lor is an allele of tbx2b/fby (from beyond). fby was identified by a pineal complex phenotype, and carries a nonsense mutation in the T-box domain of the tbx2b transcription factor. Homozygous fby mutant larvae and lor/fby transheterozygotes also display the lots-of-rods phenotype. Based upon these data, we propose a previously undescribed function for tbx2b in photoreceptor cell precursors, to promote the UV cone fate by repressing the rod differentiation pathway.

Bucky Ball Organizes Germ Plasm Assembly in Zebrafish

Current Biology : CB. Mar, 2009  |  Pubmed ID: 19249209

In many animals, gamete formation during embryogenesis is specified by maternal cytoplasmic determinants termed germ plasm. During oogenesis, germ plasm forms a distinct cellular structure such as pole plasm in Drosophila or the Balbiani body, an aggregate of organelles also found in mammals. However, in vertebrates, the key regulators of germ plasm assembly are largely unknown. Here, we show that, at the beginning of zebrafish oogenesis, the germ plasm defect in bucky ball (buc) mutants precedes the loss of polarity, indicating that Buc primarily controls Balbiani body formation. Moreover, we molecularly identify the buc gene, which is exclusively expressed in the ovary with a novel, dynamic mRNA localization pattern first detectable within the Balbiani body. We find that a Buc-GFP fusion localizes to the Balbiani body during oogenesis and with the germ plasm during early embryogenesis, consistent with a role in germ plasm formation. Interestingly, overexpression of buc seems to generate ectopic germ cells in the zebrafish embryo. Because we discovered buc homologs in many vertebrate genomes, including mammals, these results identify buc as the first gene necessary and sufficient for germ plasm organization in vertebrates.

Microtubule Actin Crosslinking Factor 1 Regulates the Balbiani Body and Animal-vegetal Polarity of the Zebrafish Oocyte

PLoS Genetics. Aug, 2010  |  Pubmed ID: 20808893

Although of fundamental importance in developmental biology, the genetic basis for the symmetry breaking events that polarize the vertebrate oocyte and egg are largely unknown. In vertebrates, the first morphological asymmetry in the oocyte is the Balbiani body, a highly conserved, transient structure found in vertebrates and invertebrates including Drosophila, Xenopus, human, and mouse. We report the identification of the zebrafish magellan (mgn) mutant, which exhibits a novel enlarged Balbiani body phenotype and a disruption of oocyte polarity. To determine the molecular identity of the mgn gene, we positionally cloned the gene, employing a novel DNA capture method to target region-specific genomic DNA of 600 kb for massively parallel sequencing. Using this technique, we were able to enrich for the genomic region linked to our mutation within one week and then identify the mutation in mgn using massively parallel sequencing. This is one of the first successful uses of genomic DNA enrichment combined with massively parallel sequencing to determine the molecular identity of a gene associated with a mutant phenotype. We anticipate that the combination of these technologies will have wide applicability for the efficient identification of mutant genes in all organisms. We identified the mutation in mgn as a deletion in the coding sequence of the zebrafish microtubule actin crosslinking factor 1 (macf1) gene. macf1 is a member of the highly conserved spectraplakin family of cytoskeletal linker proteins, which play diverse roles in polarized cells such as neurons, muscle cells, and epithelial cells. In mgn mutants, the oocyte nucleus is mislocalized; and the Balbiani body, localized mRNAs, and organelles are absent from the periphery of the oocyte, consistent with a function for macf1 in nuclear anchoring and cortical localization. These data provide the first evidence for a role for spectraplakins in polarization of the vertebrate oocyte and egg.

'ZP Domain' of Human Zona Pellucida Glycoprotein-1 Binds to Human Spermatozoa and Induces Acrosomal Exocytosis

Reproductive Biology and Endocrinology : RB&E. 2010  |  Pubmed ID: 20831819

The human egg coat, zona pellucida (ZP), is composed of four glycoproteins designated as zona pellucida glycoprotein-1 (ZP1), -2 (ZP2), -3 (ZP3) and -4 (ZP4) respectively. The zona proteins possess the archetypal 'ZP domain', a signature domain comprised of approximately 260 amino acid (aa) residues. In the present manuscript, attempts have been made to delineate the functional significance of the 'ZP domain' module of human ZP1, corresponding to 273-551 aa fragment of human ZP1.

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