NHE3 phosphorylation via PKCη marks the polarity and orientation of directionally migrating cells

Endogenous electric fields (EF) may provide an overriding cue for directional cell migration during wound closure. Perceiving a constant direction requires active sodium-hydrogen exchanger (pNHE3) at the leading edge of HEK 293 cells but its activation mechanism is not yet fully understood. Because protein kinase C (PKC) is required in electrotaxis, we asked whether NHE3 is activated by PKC during wound healing. Using pharmacological (pseudosubstrate and edelfosine) inhibition, we showed that inhibition of PKCη isoform impairs directional cell migration in HEK 293 cells in the presence of a persistent directional cue (0.25–0.3 V/mm of EF for 2 h). Further, we found that pNHE3 forms complexes with both PKCη and ?-tubulin, suggesting that these molecules may regulate the microtubule-organizing center. In addition, cellular pNHE3 content was reduced significantly when PKCη was inhibited during directional cell migration. Taken together, these data suggest that PKCη-dependent phosphorylation of NHE3 and the formation of pNHE3/PKCη/?-tubulin complexes at the leading edge of the cell are required for directional cell migration in an EF.     

Mass expulsion of zooxanthellae by heat-stressed reef corals: a source of food for giant clams?

Summary Symbiontic zooxanthellae found in the stomach of the giant clamTridacna gigas are of exogenous origin. They become available to the clam following their mass expulsion from heat-stressed hermatypic corals. The frequent appearance of these mini-plankton blooms also permits the primary production of zooxanthellae to become an available food source for other filter-feeding reef organisms rather than remaining imprisoned within the tissues of corals.We are grateful to personnel of the Mid-Pacific Marine Laboratory, Enewetak Atoll, Marshall Islands, and Friday Harbor Laboratories Washington, for providing services and facilities where portions of this study were performed. Financial support was provided by the US Department of Energy and by operating grants from the National Research Council of Canada. We thank Drs Eugene N. Kozloff and Richard R. Strathmann for comments.     

Frequent somatic mutations in MAP3K5 and MAP3K9 in metastatic melanoma identified by exome sequencing

We sequenced eight melanoma exomes to identify new somatic mutations in metastatic melanoma. Focusing on the mitogen-activated protein (MAP) kinase kinase kinase (MAP3K) family, we found that 24% of melanoma cell lines have mutations in the protein-coding regions of either MAP3K5 or MAP3K9. Structural modeling predicted that mutations in the kinase domain may affect the activity and regulation of these protein kinases. The position of the mutations and the loss of heterozygosity of MAP3K5 and MAP3K9 in 85% and 67% of melanoma samples, respectively, together suggest that the mutations are likely to be inactivating. In in vitro kinase assays, MAP3K5 I780F and MAP3K9 W333* variants had reduced kinase activity. Overexpression of MAP3K5 or MAP3K9 mutants in HEK293T cells reduced the phosphorylation of downstream MAP kinases. Attenuation of MAP3K9 function in melanoma cells using siRNA led to increased cell viability after temozolomide treatment, suggesting that decreased MAP3K pathway activity can lead to chemoresistance in melanoma.     

Patterns and rates of exonic de novo mutations in autism spectrum disorders

Autism spectrum disorders (ASD) are believed to have genetic and environmental origins, yet in only a modest fraction of individuals can specific causes be identified. To identify further genetic risk factors, here we assess the role of de novo mutations in ASD by sequencing the exomes of ASD cases and their parents (n = 175 trios). Fewer than half of the cases (46.3%) carry a missense or nonsense de novo variant, and the overall rate of mutation is only modestly higher than the expected rate. In contrast, the proteins encoded by genes that harboured de novo missense or nonsense mutations showed a higher degree of connectivity among themselves and to previous ASD genes as indexed by protein-protein interaction screens. The small increase in the rate of de novo events, when taken together with the protein interaction results, are consistent with an important but limited role for de novo point mutations in ASD, similar to that documented for de novo copy number variants. Genetic models incorporating these data indicate that most of the observed de novo events are unconnected to ASD; those that do confer risk are distributed across many genes and are incompletely penetrant (that is, not necessarily sufficient for disease). Our results support polygenic models in which spontaneous coding mutations in any of a large number of genes increases risk by 5- to 20-fold. Despite the challenge posed by such models, results from de novo events and a large parallel case-control study provide strong evidence in favour of CHD8 and KATNAL2 as genuine autism risk factors.     

Electrical signals control wound healing through phosphatidylinositol-3-OH kinase-gamma and PTEN

Wound healing is essential for maintaining the integrity of multicellular organisms. In every species studied, disruption of an epithelial layer instantaneously generates endogenous electric fields, which have been proposed to be important in wound healing. The identity of signalling pathways that guide both cell migration to electric cues and electric-field-induced wound healing have not been elucidated at a genetic level. Here we show that electric fields, of a strength equal to those detected endogenously, direct cell migration during wound healing as a prime directional cue. Manipulation of endogenous wound electric fields affects wound healing in vivo. Electric stimulation triggers activation of Src and inositol-phospholipid signalling, which polarizes in the direction of cell migration. Notably, genetic disruption of phosphatidylinositol-3-OH kinase-gamma (PI(3)Kgamma) decreases electric-field-induced signalling and abolishes directed movements of healing epithelium in response to electric signals. Deletion of the tumour suppressor phosphatase and tensin homolog (PTEN) enhances signalling and electrotactic responses. These data identify genes essential for electrical-signal-induced wound healing and show that PI(3)Kgamma and PTEN control electrotaxis.