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Alternative polyadenylation (APA) is an important contributor to the regulation of gene expression in plants. One subunit of the complex that cleaves and polyadenylates mRNAs in the nucleus, CPSF30 (for the 30 kD subunit of the mammalian Cleavage and Polyadenylation Specificity Factor), has been implicated in a wide-ranging network of regulatory events. CPSF30 plays roles in root development, flowering time, and response to biotic and abiotic stresses. CPSF30 also is a conduit that links cellular signaling and RNA modification with alternative RNA processing events and transcriptional dynamics. While much is known about CPSF30 and its roles in plants, questions remain regarding the connections between CPSF30-mediated APA and the downstream events that lead to specific phenotypic outcomes. To address these, we conducted a detailed analysis of poly(A) site usage in the CPSF30 mutant. Our results corroborate earlier reports that link CPSF30 with a distinctive cis element (AAUAAA) that is present 10-30 nts upstream of some, but not all, plant pre-mRNAs. Interestingly, our results reveal a distinctive shift in poly(A) site in mutants deficient in CPSF30, resulting in cleavage and polyadenylation at the location of motifs similar to AAUAAA. Importantly, CPSF30-associated APA had at best a small impact on mRNA functionality. These results necessitate the formulation of new hypotheses for mechanisms by which CPSF30-mediated APA influences physiological processes. 

The cytokinetic contractile ring (CR) was first described some 50 years ago, however our understanding of the assembly and structure of the animal cell CR remains incomplete. We recently reported that mature CRs in sea urchin embryos contain myosin II mini-filaments organized into aligned concatenated arrays, and that in early CRs myosin II formed discrete clusters that transformed into the linearized structure over time. The present study extends our previous work by addressing the hypothesis that these myosin II clusters also contain the crucial scaffolding proteins anillin and septin, known to help link actin, myosin II, RhoA, and the membrane during cytokinesis. Super-resolution imaging of cortices from dividing embryos indicates that within each cluster, anillin and septin2 occupy a centralized position relative to the myosin II mini-filaments. As CR formation progresses, the myosin II, septin and anillin containing clusters enlarge and coalesce into patchy and faintly linear patterns. Our super-resolution images provide the initial visualization of anillin and septin nanostructure within an animal cell CR, including evidence of a septin filament-like network. Furthermore, Latrunculin-treated embryos indicated that the localization of septin or anillin to the myosin II clusters in the early CR was not dependent on actin filaments. These results highlight the structural progression of the CR in sea urchin embryos from an array of clusters to a linearized purse string, the association of anillin and septin with this process, and provide the visualization of an apparent septin filament network with the CR structure of an animal cell.