In sexual reproduction of most animals, the spermatozoon provides DNA and centrioles, together with some cytoplasm and organelles, to the oocyte that is being fertilized. Paternal mitochondria and their genomes are generally eliminated in the embryo by an unknown degradation mechanism. We show that, upon fertilization, a Caenorhabditis elegans spermatozoon triggers the recruitment of autophagosomes within minutes and subsequent paternal mitochondria degradation. Whereas the nematode-specific sperm membranous organelles are ubiquitinated before autophagosome formation, the mitochondria are not. The degradation of both paternal structures and mitochondrial DNA requires an LC3-dependent autophagy. Analysis of fertilized mouse embryos shows the localization of autophagy markers, which suggests that this autophagy event is evolutionarily conserved to prevent both the transmission of paternal mitochondrial DNA to the offspring and the establishment of heteroplasmy.

Asymmetric localization of PAR proteins is a hallmark of polarized cells, but the mechanisms that create PAR asymmetry are not well understood. In the C. elegans zygote, PAR asymmetry is initiated by a transient actomyosin contraction, which sweeps the PAR-3/PAR-6/PKC-3 complex toward the anterior pole of the egg. The RING finger protein PAR-2 accumulates in a complementary pattern in the posterior cortex. Here we present evidence that PAR-2 participates in a feedback loop to stabilize polarity. PAR-2 is a target of the PKC-3 kinase and is excluded from the anterior cortex by PKC-3-dependent phosphorylation. The RING domain of PAR-2 is required to overcome inhibition by PKC-3 and stabilize PAR-2 on the posterior cortex. Cortical PAR-2 in turn prevents PAR-3/PAR-6/PKC-3 from returning to the posterior, in a PAR-1- and PAR-5-dependent manner. Our findings suggest that reciprocal inhibitory interactions among PAR proteins stabilize polarity by reinforcing an initial asymmetry in PKC-3.

The par-2 gene of Caenorhabditi elgans functions in early embryogenesis to ensure an amme first cleavage and the segregation of cytoplasmic factors. Both processes appear to be required to generate daughter blastomeres with distinct developmental potential. We isolated an allele of par-2 by using a screen for maternal-effect lethal mutations in a strain known for its high frequency of transposition events. A transposable lement was found to be linked to this allele. Sequences lanig the site oftransposon insertion were cloned and found to rescue the par-2 mutant phenotype. DNA In the par-2 region hybridized to a 2.3-kb germ-liueenriched mRNA. The cDNA corresponding to this germ-lineenriched message was cloned, sequenced, and used to identify the molecular lesions associated with three par-2 alleles. Sequence analysis of the par-2 cDNA revealed that the predicted protein contained two distinct motifs found in other known proteins: an ATP-binding site and a cystelue-rich motif which identifies thepar-2 gene product as a member of a growing class of putative zinc-binding proteins.A class of genes has been identified that is required for the generation of asymmetry in the early embryo and whose mutant phenotype resembles the cytochalasin D-treated embryos described above (6). Mutations in any one of the five par genes (for partitioning-defective) result in embryos with defects in the pition of the first cleavage furrow, altered orientation of the spindle at the second cleavage, synchronous early cleavages, defects in P-granule localization, and altered expression of intestinal cell differentiation markers. All mutations in these five genes result in strict maternaleffect lethality; therefore maternal expression of the par genes is required during oogenesis in order to produce viable embryos.To gain further insight into the role of the par genes in cytoplasmic partitioning, molecular analysis ofthese genes is essential. As a step toward an understanding of the generation of asymmetry and cytoplasmic localization in the C. elegans embryo at the molecular level, we have isolated and sequenced the par-2 gene. 1 1 Early embryogenesis in the nematode Caenorhabditis elegans is characterized by a series of asymmetric cleavages. The initial asymmetry of the early embryo becomes apparent during the first cell cycle, at which time cytoplasmic factors thought to be required for the specification of cell fate are segregated. The first cleavage produces a larger, anterior cell (AB) and a smaller, posterior cell (P1) which differ from each other in many respects including the orientation of subsequent cleavage planes, the timing ofcell cycles, and the kinds of differentiated cell types they ultimately produce (1). For instance, germ-line-specific P granules are uniformly dispersed in one-cell embryos but become localized to the posterior cortex midway through the first cell cycle (2). At first cleavage, nearly all of the P granules are partitioned to the P1 blastomere. Furthermore, studies of gut development (3) have shown ...

Background: Protein aggregation is a hallmark of several neurodegenerative diseases including Huntington's disease and Parkinson's disease. Proteins containing long, homopolymeric stretches of glutamine are especially prone to form aggregates. It has long been known that the small protein modifier, ubiquitin, localizes to these aggregates. In this report, nematode and cell culture models for polyglutamine aggregation are used to investigate the role of the ubiquitin pathway in protein aggregation.

The RING finger motif exists in E3 ligases of the ubiquitination pathway. These ubiquitin ligases bind to target proteins, leading to their modification by covalent addition of ubiquitin peptides. Current databases contain hundreds of proteins with RING finger motifs. This study investigates the role of RING finger genes in embryogenesis of the nematode, Caenorhabditis elegans. We expand the previous list of RING finger-containing genes and show that there are 103 RING finger-containing genes in the C. elegans genome. DNA microarrays of these 103 genes were probed with various RNA samples to identify 16 RING finger genes whose expression is enriched in the germline. RNA interference (RNAi) analysis was then used to determine the developmental role of these genes. One RING finger gene, C32D5.10, showed a dramatic larval arrest upon RNAi. Three RING finger genes exhibited embryonic lethality after RNAi. These three genes include par-2, and two small RING finger proteins: F35G12.9 (an ortholog of APC11) and ZK287.5 (an ortholog of rbx1). Embryos from RNAi of the APC11 and rbx1 orthologs were arrested in the cell cycle, confirming the role of these particular RING finger proteins in regulation of the cell cycle. genesis 38:1-12, 2004.