Analysis of Factors that Regulate Asymmetric Cell Division in the Green Alga Volvox Carteri

Analysis of Factors that Regulate Asymmetric Cell Division in the Green Alga Volvox Carteri

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Asymmetric cell divisions are critical for generating cellular diversity in multicellular organisms. Volvox carteri is an excellent model system for the study of asymmetric cell division as it possesses only two cell types (aˆ¼16 large reproductive cells called gonidia and aˆ¼2000 small, motile somatic cells) whose progenitors are set apart by asymmetric cell divisions early in embryogenesis. Mutations at gls (gonidialess) loci selectively abolish asymmetric divisions and cause a Gls (Gonidialess) phenotype in which all cells are small and develop as somatic cells. One gls gene, glsA, has been cloned. It encodes a J protein chaperone (GlsA) with four evolutionarily conserved domains, named J, M, SANT1, and SANT2. To learn more about GlsA function I generated derivatives of GlsA that are mutated in its conserved domains, and I carried out a yeast two-hybrid interaction screen to identify binding partners of GlsA. I found that all of the conserved domains of GlsA are necessary for its asymmetric division function but that the SANT domains are not essential for a newly identified function that probably involves translation, suggesting that GlsA does not regulate asymmetric division through this translation function. In addition, GlsA and derivatives that contains the SANT2 domain co-immunoprecipitated with histones, suggesting that GlsA likely functions at some level in transcription, and that the SANT2 domain is required for this function. One of a small number of polypeptides that reproducibly interacted with GlsA in my yeast two-hybrid screen was a aˆ¼200-aa segment of a protein homologous to the evolutionarily conserved DP proteins, which are components of the retinoblastoma protein pathway that regulates the cell cycle and cell differentiation in plants and animals. I cloned this DP-encoding gene (dp1) and characterized it and its protein product, DP1. Two DP1 protein isoforms could be detected in cell extracts made from a transgenic strain expressing myc-tagged DP1, one that migrated at aˆ¼60 kd on SDS-PAGE and accumulated only in somatic cells, and one that migrated at aˆ¼80 kd and accumulated only in reproductive cells and young juveniles. Both isoforms co-immunoprecipitated with GlsA, and DP1 and GlsA both co-localize with chromatin, indicating that DP1 and GlsA are very likely to belong to the same complex in V. carteri embryos. Expression of a version of DP1 that was expected to act as a dominant negative against endogenous DP1 function did not lead to an asymmetric cell division phenotype in a V. carteri strain expressing the transcript for the protein from its native promoter. Therefore while it is likely that GlsA and DP1 belong to the same transcriptional complex, it is not clear, yet, whether any of the targets of this complex are genes required for asymmetric division.Gene diagram key is as described in Figure 4.2C, except for following. The short vertical line above the third exon indicates position of c-myc tag, and the unfilled box labeled a€œaˆ†a€ indicates the region of the gene that encodes the first Ip helix ofanbsp;...

Title:Analysis of Factors that Regulate Asymmetric Cell Division in the Green Alga Volvox Carteri
Author: Valeria Pappas
Publisher:ProQuest - 2008

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