Collaboration within the M1 aminopeptidase family promotes reproductive success in Caenorhabditis elegans.

Mutations of the puromycin-sensitive aminopeptidase (Psa) orthologs of flies, mice, and plants result in meiotic errors and reduced embryonic viability. Genetic lesions of the Caenorhabditis elegans ortholog of Psa, pam-1, similarly result in dramatic reductions of worm fecundity. The gonads of animals harboring mutant pam-1 alleles display expanded populations of pachytene germinal nuclei and delayed nucleolar disassembly in the developing oocytes, phenotypes that ultimately hinder embryonic viability and overall brood sizes. PAM-1 is a member of the M1 aminopeptidase family and shares a high amount of homology with its M1 paralogs. Comparative analysis of the M1 aminopeptidase family reveals that only nine (including PAM-1) of the 17 annotated M1 aminopeptidases are predicted to be catalytically active. Interestingly, we demonstrate that three of these active M1 paralogs have roles independent of PAM-1 in promoting gametogenesis and fecundity. Simultaneous inhibition of pam-1 and M1 paralogs produces synergistic decreases in overall brood sizes and embryonic viability, exacerbates the germinal phenotypes of pachytene extension and delayed nucleolar disassembly, and unmasks previously hidden phenotypes. Our data suggests that the interdependent functions of multiple M1 aminopeptidases are necessary for reproductive success in C. elegans and lend further credence to the redundant composition of an evolutionarily conserved enzyme family.

Wnt signaling drives WRM-1/beta-catenin asymmetries in early C. elegans embryos.

beta-Catenin regulates cell adhesion and cellular differentiation during development, and misregulation of beta-catenin contributes to numerous forms of cancer in humans. Here we describe Caenorhabditis elegans conditional alleles of mom-2/Wnt, mom-4/Tak1, and wrm-1/beta-catenin. We use these reagents to examine the regulation of WRM-1/beta-catenin during a Wnt-signaling-induced asymmetric cell division. While WRM-1 protein initially accumulates in the nuclei of all cells, signaling promotes the retention of WRM-1 in nuclei of responding cells. We show that both PRY-1/Axin and the nuclear exportin homolog IMB-4/CRM-1 antagonize signaling. These findings reveal how Wnt signals direct the asymmetric localization of beta-catenin during polarized cell division.

The minibrain kinase homolog, mbk-2, is required for spindle positioning and asymmetric cell division in early C. elegans embryos.

In the newly fertilized Caenorhabditis elegans zygote, cytoplasmic determinants become localized asymmetrically along the anterior-posterior (A-P) axis of the embryo. The mitotic apparatus then orients so as to cleave the embryo into anterior and posterior blastomeres that differ in both size and developmental potential. Here we describe a role for MBK-2, a member of the Dyrk family of protein kinases, in asymmetric cell division in C. elegans. In mbk-2 mutants, the initial mitotic spindle is misplaced and cytoplasmic factors, including the germline-specific protein PIE-1, are mislocalized. Our findings support a model in which MBK-2 down-regulates the katanin-related protein MEI-1 to control spindle positioning and acts through distinct, as yet unknown factors, to control the localization of cytoplasmic determinants. These findings in conjunction with work from Schizosaccharomyces pombe indicate a possible conserved role for Dyrk family kinases in the regulation of spindle placement during cell division.

Attenuated APC alleles produce functional protein from internal translation initiation.

Some truncating mutations of the APC tumor suppressor gene are associated with an attenuated phenotype of familial adenomatous polyposis coli (AAPC). This work demonstrates that APC alleles with 5' mutations produce APC protein that down-regulates beta-catenin, inhibits beta-catenin/T cell factor-mediated transactivation, and induces cell-cycle arrest. Transfection studies demonstrate that cap-independent translation is initiated internally at an AUG at codon 184 of APC. Furthermore, APC coding sequence between AAPC mutations and AUG 184 permits internal ribosome entry in a bicistronic vector. These data suggest that AAPC alleles in vivo may produce functional APC by internal initiation and establish a functional correlation between 5' APC mutations and their associated clinical phenotype.