SUMMARY Golgi membranes, from yeast to humans, are uniquely enriched in phosphatidylinositol-4-phosphate (PtdIns(4)P), although the role of this lipid remains poorly understood. Using a proteomic lipid binding screen, we identify the Golgi protein GOLPH3 (also called GPP34, GMx33, MIDAS, or yeast Vps74p) as a PtdIns(4)P-binding protein that depends upon PtdIns(4)P for its Golgi localization. We further show that GOLPH3 binds the unconventional myosin MYO18A, thus connecting the Golgi to F-actin. We demonstrate that this linkage is necessary for normal Golgi trafficking and morphology. The evidence suggests that GOLPH3 binds to PtdIns(4)P-rich trans-Golgi membranes and MYO18A conveying a tensile force required for efficient tubule and vesicle formation. Consequently, this tensile force stretches the Golgi into the extended ribbon observed by fluorescence microscopy and the familiar flattened form observed by electron microscopy.

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Many axons carry out the synthesis of macromolecules independent of their cell bodies but the nature, organization and magnitude of axonal protein synthesis remain unclear. We have examined these features in axons of chick sympathetic neurons in cell culture. In situ hybridization showed that poly(A) mRNA is abundant and non-uniformly distributed in nearly all axons. The specific transcripts for β-actin and actin-depolymerizing factor (ADF) were also present and non-uniformly distributed in axons, with an approximately hundredfold higher concentration in growth cones, branch points and axonal varicosities than in the axon shaft. Immunoprecipitation using specific antibodies indicates that β-actin, ADF and neurofilament protein (NF) are translated in axons independently of cell bodies. Quantification of the distribution of β-actin and ADF mRNAs showed that their ability to enter the axon was likely to be a property of the neuron as a whole rather than of individual axons. To compare the distribution of axonally translated protein to that of mRNA, we performed 35S metabolic labeling with axons separated from their cell bodies. Axonally synthesized proteins were distributed throughout the axons and their synthesis was inhibited by cycloheximide but not by chloramphenicol. Proteins translated mainly or exclusively in axons or cell bodies were both detected by metabolic labeling. Axons separated from their cell bodies synthesized up to 5% as much protein in a 3-hour period as did intact neurons. Because axons in our culture conditions contain ∼50% of the non-nuclear volume of the neurons, we estimate that axoplasm of sympathetic neurons has a protein synthetic capacity per unit volume equal to 10% that of cell body cytoplasm.

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Caenorhabditis elegans is a free living soil nematode and thus in its natural habitat, C. elegans encounters many different species of soil bacteria. Although some soil bacteria may be excellent sources of nutrition for the worm, others may be pathogenic. Thus, we undertook a study to understand how C. elegans can identify their preferred food using a simple behavioral assay. We found that there are various species of soil bacteria that C. elegans prefers in comparison to the standard laboratory E. coli strain OP50. In particular, two bacterial strains, Bacillus mycoides and Bacillus soli, were preferred strains. Interestingly, the sole feeding of these bacteria to wild type animals results in extended lifespan through the activation of the autophagic process. Further studies will be required to understand the precise mechanism controlling the behavior of identification and selection of food in C. elegans.

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We demonstrated that nicotine inhibits growth through cell cycle arrest at G(0)/G(1) phase probably by increasing the expression of p21(WAF1/CIP1). Nicotine also affects epithelial differentiation in immortalized and malignant oral keratinocytes. Malignant oral keratinocytes appear to be more resistant to the effects of nicotine on epithelial growth and differentiation as compared to the immortalized cells.