Functional redundancy of two C. elegans homologs of the histone chaperone Asf1 in germline DNA replication.

Eukaryotic genomes contain either one or two genes encoding homologs of the highly conserved histone chaperone Asf1, however, little is known of their in vivo roles in animal development. UNC-85 is one of the two Caenorhabditis elegans Asf1 homologs and functions in post-embryonic replication in neuroblasts. Although UNC-85 is broadly expressed in replicating cells, the specificity of the mutant phenotype suggested possible redundancy with the second C. elegans Asf1 homolog, ASFL-1. The asfl-1 mRNA is expressed in the meiotic region of the germline, and mutants in either Asf1 genes have reduced brood sizes and low penetrance defects in gametogenesis. The asfl-1, unc-85 double mutants are sterile, displaying defects in oogenesis and spermatogenesis, and analysis of DNA synthesis revealed that DNA replication in the germline is blocked. Analysis of somatic phenotypes previously observed in unc-85 mutants revealed that they are neither observed in asfl-1 mutants, nor enhanced in the double mutants, with the exception of enhanced male tail abnormalities in the double mutants. These results suggest that the two Asf1 homologs have partially overlapping functions in the germline, while UNC-85 is primarily responsible for several Asf1 functions in somatic cells, and is more generally involved in replication throughout development.

UNC-85, a C. elegans homolog of the histone chaperone Asf1, functions in post-embryonic neuroblast replication.

Normal animal development requires accurate cell divisions, not only in the early stages of rapid embryonic cleavages, but also in later developmental stages. The Caenorhabditis elegans unc-85 gene is implicated only in cell divisions that occur post-embryonically, primarily in terminal neuronal lineages. Variable post-embryonic cell division failures in ventral cord motoneuron precursors result in uncoordinated locomotion of unc-85 mutant larvae by the second larval stage. These neuroblast cell division failures often result in unequally sized daughter nuclei, and sometimes in nuclear fusions. Using a combination of conventional mapping techniques and microarray analysis, we cloned the unc-85 gene, and find that it encodes one of two C. elegans homologs of the yeast Anti-silencing function 1 (Asf1) histone chaperone. The unc-85 gene is expressed in replicating cells throughout development, and the protein is localized in nuclei. Examination of null mutants confirms that embryonic neuroblast cell divisions occur normally, but post-embryonic neuroblast cell divisions fail. Analysis of the DNA content of the mutant neurons indicates that defective replication in post-embryonic neuroblasts gives rise to ventral cord neurons with an average DNA content of approximately 2.5 n. We conclude that UNC-85 functions in post-embryonic DNA replication in ventral cord motor neuron precursors.

A computational model for C. elegans locomotory behavior: application to multiworm tracking.

A computational approach is presented for modeling and quantifying the structure and dynamics of the nematode C. elegans observed by time-lapse microscopy. Worm shape and conformations are expressed in a decoupled manner. Complex worm movements are expressed in terms of three primitive patterns--peristaltic progression, deformation, and translation. The model has been incorporated into algorithms for segmentation and simultaneous tracking of multiple worms in a field, some of which may be interacting in complex ways. A recursive Bayesian filter is used for tracking. Unpredictable behaviors associated with interactions are resolved by multiple-hypothesis tracking. Our algorithm can track worms of diverse sizes and conformations (coiled/uncoiled) in the presence of imaging artifacts and clutter, even when worms are overlapping with others. A two-observer performance assessment was conducted over 16 image sequences representing wild-type and uncoordinated mutants as a function of worm size, conformation, presence of clutter, and worm entanglement. Overall detected tracking failures were 1.41%, undetected tracking failures were 0.41%, and segmentation errors were 1.11% of worm length. When worms overlap, our method reduced undetected failures from 12% to 1.75%, and segmentation error from 11% to 5%. Our method provides the basis for reliable morphometric and locomotory analysis of freely behaving worm populations.

Reining in cytokinesis with a septin corral.

Septins are a family of conserved GTP-binding proteins that function in cytokinesis in fungi and animals. In budding yeast, septins form scaffolds for assembly of the actomyosin contractile ring at the cleavage plane, a role that does not appear to be conserved in other organisms. The septins form an hourglass-shaped collar at the mother-bud neck, which splits into two rings flanking the division plane at cytokinesis. A recent study(1) demonstrates that these two septin rings constitute diffusion barriers that create a cytokinetic compartment to retain cortical cytokinetic factors in proximity to the cleavage plane.

Interdependence of the contractile ring and spindle midzone in cleavage plane maintenance.

How segregation of the chromosomes is coordinated with the ensuing cell cleavage to complete the cell cycle is not well understood. A recent study of cytokinesis in fission yeast by Pardo and Nurse suggests that the contractile ring is required for assembly of the post-mitotic microtubule array (PAA). In turn, the PAA is required to maintain the contractile ring at the cleavage plane, as well as to keep the nuclei separated at the poles of the cleaving cell. These functions may be particularly important for a cell cycle checkpoint ensuring that if cytokinesis is delayed, septation will occur between the two daughter nuclei.

A role for septins in cellular and axonal migration in C. elegans.

Caenorhabditis elegans has two genes, unc-59 and unc-61, encoding septin-family GTPases. Mutations in the septin genes cause defects in locomotory behavior that have been previously attributed to cytokinesis failures in postembryonic neuroblasts. We find that mutations in either septin gene frequently cause uncoordination in newly hatched larvae in the absence of cytokinesis failures. The septins exhibit developmentally regulated expression, including expression in various neurons at times when processes are extending and synapses are forming. Motor neurons in the mutant larvae display defects in multiple aspects of axonal migration and guidance that are likely to be responsible for the locomotory behavior defects. The septins are also expressed in migrating distal tip cells, which are leaders for gonad arm extension. Septin mutants affect morphology of the distal tip cells, as well as their migration and guidance during gonadogenesis. These results suggest that septins may be generally required for developmental migrations and pathfinding.

One ring to bind them. Septins and actin assembly.

Septins are GTPases required for cytokinesis and other processes requiring spatial organization of the cell cortex, but their molecular functions in these processes are unknown. In this issue of Developmental Cell, Kinoshita et al. take an important step in elucidating the molecular functions of septins by developing an in vitro assay for septin assembly and exploring the relationship between mammalian septins and actin.

Fusion and fission: membrane trafficking in animal cytokinesis.

Cytokinesis is the physical act of separating daughter cells, allowing them to become separate entities. Recent studies have revealed that membrane insertion for furrowing and scission of the residual bridge is a key aspect of animal cytokinesis.