The nematode Caenorhabditis elegans uses striated muscle in its body wall for locomotion. The myofilament lattice is organized such that all the thin filament attachment structures (dense bodies, analogous to Z-disks) and thick filament organizing centers (M-lines) are attached to the muscle cell membrane. Thus, the force of muscle contraction is transmitted through these structures and allows locomotion of the worm. Dense bodies and M-lines are compositionally similar to focal adhesions and costameres, and are based on integrin and associated proteins. Null mutants for many of the newly discovered dense body and M-line proteins do not have obvious locomotion defects when observed casually, or when assayed by counting the number of times a worm moves back and forth in liquid. We hypothesized that many of these proteins, located as they are in muscle focal adhesions, function in force transmission, but we had not used an appropriate or sufficiently sensitive assay to reveal this function. Recently, we have developed a new quantitative assay of C. elegans locomotion that measures the maximum bending amplitude of an adult worm as it moves backwards. The assay had been used to reveal locomotion defects for null mutants of genes encoding ATN-1 (α-actinin) and PKN-1 (protein kinase N). Here, we describe the details of this method, and apply it to 21 loss of function mutants in 17 additional genes, most of which encode components of muscle attachment structures. As compared to wild type, mutations in 11 genes were found to have less ability to bend, and mutations in one gene were found to have greater ability to bend. Loss of function mutants for eight proteins had been reported to have normal locomotion (ZYX-1(zyxin), ALP-1 (Enigma), DIM-1, SCPL-1), or locomotion that was not previously investigated (FRG-1 (FRG1), KIN-32 (focal adhesion kinase, LIM-8), or had only slightly decreased locomotion (PFN-3 (profilin)).

Studying HIV-1 replication in the presence of functionally related proteins from different species has helped define host determinants of HIV-1 infection. Humans and owl monkeys, but not macaques, encode a CD4 receptor that permits entry of transmissible HIV-1 variants due to a single residue difference. However, little is known about whether divergent CCR5 receptor proteins act as determinants of host-range. Here we show that both owl monkey (Aotus vociferans) CD4 and CCR5 receptors are functional for the entry of transmitted HIV-1 when paired with human versions of the other receptor. By contrast, the owl monkey CD4/CCR5 pair is generally a suboptimal receptor combination, although there is virus-specific variation in infection with owl monkey receptors. Introduction of the human residues 15Y and 16T within a sulfation motif into owl monkey CCR5 resulted in a gain of function. These findings suggest there is cross-talk between CD4 and CCR5 involving the sulfation motif.

To examine the in vivo functions of protein kinase N (PKN), one of the effectors of Rho small GTPases, we used the nematode Caernorhabditis elegans as a genetic model system. We identified a C. elegans homologue (pkn-1) of mammalian PKN and confirmed direct binding to C. elegans Rho small GTPases. Using a GFP reporter, we showed that pkn-1 is mainly expressed in various muscles and is localized at dense bodies and M-lines. Over-expression of the PKN-1 kinase domain and loss-of-function mutations by genomic deletion of pkn-1 resulted in a loopy Unc phenotype, which has been reported in many mutants of neuronal genes. The results of mosaic analysis and body wall muscle specific expression of PKN-1 kinase domain suggests that this loopy phenotype is due to the expression of PKN-1 in body wall muscle. The genomic deletion of pkn-1 also showed a defect in force transmission. These results suggest that PKN-1 functions as a regulator of muscle contraction-relaxation and as a component of the force transmission mechanism.