Biomedical subjects

J A Schisa

Publications and source records attributed to J A Schisa.

5 recordsLinked to original sources

PubMed2001

Analysis of RNA associated with P granules in germ cells of C. elegans adults.

P granules are cytoplasmic structures of unknown function that are associated with germ nuclei in the C. elegans gonad, and are localized exclusively to germ cells, or germ cell precursors, throughout the life cycle. All the known protein components of P granules contain putative RNA-binding motifs, suggesting that RNA is involved in either the structure or function of the granules. However, no specific mRNAs have been identified within P granules in the gonad. We show here that P granules normally contain a low level of RNA, and describe conditions that increase this level. We present evidence that several, diverse mRNAs, including pos-1, mex-1, par-3, skn-1, nos-2 and gld-1 mRNA, are present at least transiently within P granules. In contrast, actin and tubulin mRNA and rRNA are either not present in P granules, or are present at relatively low levels. We show that pgl-1 and the glh (Vasa-related) gene family, which encode protein components of P granules, do not appear essential for RNA to concentrate in P granules; these proteins may instead function in events that are a prerequisite for RNAs to be transported efficiently from the nuclear surface.

Animals↗

PubMed2000

P granules in the germ cells of Caenorhabditis elegans adults are associated with clusters of nuclear pores and contain RNA.

The germ cells, and germ cell precursors, in the nematode Caenorhabditis elegans contain distinctive granules called P granules. During early embryogenesis, P granules are segregated asymmetrically into those blastomeres that eventually produce the germ line. Because of the correlation between P granule distribution and the development of the germ line, P granules are widely thought to function in some aspect of germ line specification or differentiation. Most of the analysis of P granule structure and localization has focused on the early embryo, when P granules are located in the cytoplasm. However, during most of development P granules are associated with germ cell nuclei. We report here an ultrastructural analysis of the nuclear-associated P granules in the germ cells of the adult hermaphrodite gonad. We show that P granules are tightly associated with nuclear pores and that the positions of certain structures within the P granules correspond to the positions of pores on the nuclear envelope. We present immunocytochemical and ultrastructural data suggesting that P granules can associate, or remain associated, with pore-like structures even after they detach from the nuclear envelope during oogenesis. Finally, we show that nuclear-associated P granules in the gonad contain RNA, complementing previous studies showing that cytoplasmic P granules in embryos contain RNA.

Animals↗

PubMed1998

Cytoplasmic polyadenylation of Toll mRNA is required for dorsal-ventral patterning in Drosophila embryogenesis.

Toll encodes a receptor that is critical for dorsal-ventral patterning in the early Drosophila embryo. Previous data have suggested that the accumulation of Toll protein in the embryo temporally correlates with elongation of the poly (A) tail of the message. Here, we demonstrate that Toll mRNA is translationally activated by regulated cytoplasmic polyadenylation. We also identify a 192 nucleotide regulatory element in the Toll 3' UTR that is necessary for robust translational activation of Toll mRNA and also regulates polyadenylation. UV crosslinking analyses suggest that two proteins bind specifically to the 192 nucleotide element. One or both of these proteins may be factors that are required for translational regulation or cytoplasmic polyadenylation. These studies demonstrate that regulated polyadenylation plays a critical role in the Drosophila dorsal-ventral patterning system.

Animals↗

PubMed1997

Nanos and pumilio establish embryonic polarity in Drosophila by promoting posterior deadenylation of hunchback mRNA.

Nanos protein promotes abdominal structures in Drosophila embryos by repressing the translation of maternal hunchback mRNA in the posterior. To study the mechanism of nanos-mediated translational repression, we first examined the mechanism by which maternal hunchback mRNA is translationally activated. In the absence of nanos activity, the poly(A) tail of hunchback mRNA is elongated concomitant with its translation, suggesting that cytoplasmic polyadenylation directs activation. However, in the presence of nanos the length of the hunchback mRNA poly(A) tail is reduced. To determine if nanos activity represses translation by altering the polyadenylation state of hunchback mRNA, we injected various in vitro transcribed RNAs into Drosophila embryos and determined changes in polyadenylation. Nanos activity reduced the polyadenylation status of injected hunchback RNAs by accelerating their deadenylation. Pumilio activity, which is necessary to repress the translation of hunchback, is also needed to alter polyadenylation. An examination of translation indicates a strong correlation between poly(A) shortening and suppression of translation. These data indicate that nanos and pumilio determine posterior morphology by promoting the deadenylation of maternal hunchback mRNA, thereby repressing its translation.

Animals↗

PubMed1993

Recombinant retroviruses containing novel reporter genes.

Two histochemical marker genes, Drosophila alcohol dehydrogenase (ADH) and human placental alkaline phosphatase (ALP), were cloned into the recombinant retroviral vectors pLJ and pgag beta-actin. The resulting vectors were transfected into retroviral producer cell lines, psi CRE and psi CRIP, and stable recombinant retrovirus producers were isolated. Recombinant virus was harvested and used to transduce genes into several cell lines, singly or in conjunction with lacZ (Escherichia coli beta-galactosidase)-containing retrovirus. Cell lines were then stained using standard histochemical methods for recombinant gene expression. We found that multiple gene products could be identified in the same cell populations and in the case of ALP and beta-galactosidase, in the same cells. The resulting reagents should be useful for a variety of cell-marking studies including those involving multiple clonal analysis and developmental studies for gene therapy.

3T3 Cells↗