This is a proposal to study the cellular localization of a novel family of transcripts. Dr. Etkin's laboratory has cloned a family of genes that code for interspersed repeat transcripts from Xenopus laevis (xlsirts) and Xenopus tropicalis (xtsirts). Sirts are organized as tandem repeat sequences consisting of 79-81 nt repeats that are reiterated from 3 to 13 times in different clones. There are no open reading frames and in each clone the repeats are flanked by different unique sequences. Using in situ hybridization analysis with the repeat sequence probe the xlsirts were detected in the mitochondrial cloud of stage 1 and 2 oocytes after which they translocate to the vegetal cortical egion. The xlsirt transcripts are one of the earliest indices of animal-vegetal polarity. The hypothesis is that they may function as a structural component of the vegetal cortex and may be responsible for the subsequent localization of transcripts such as xcat2 or vg1 in the vegetal cortex.In addition, their localization in this region may influence processes such as germ cell formation or the cotical rotation which influences dorsal-ventral axis formation.
The aims are: (1) to test the role of cytoskeletal elements in the localization and anchoring of xlsirt transcripts; (2) to analyze the role of the cis-acting elements in the localization of xlsirts; (3) to analyze the role of xlsirts in the localization of other RNAs such as xcat2, xcat3, and vg1 in oocytes and to determine their function during development; and (4) to clone xlsirt-like sequences from other organisms such as Drosophila and mouse and to examine the localization of xlsirts in a heterologous system.