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Abstract. MYO2 encodes a type V myosin heavy chain needed for the targeting of vacuoles and secretory vesicles to the growing bud of yeast. Here we describe new myo2 alleles containing conditional lethal mutations in the COOH-terminal tail domain. Within 5 min of shifting to the restrictive temperature, the polarized distribution of secretory vesicles is abolished without affecting the distribution of actin or the mutant Myo2p, showing that the tail has a direct role in vesicle targeting. We also show that the actin cable–dependent translocation of Myo2p to growth sites does not require secretory vesicle cargo. Although a fusion protein containing the Myo2p tail also concentrates at growth sites, this accumulation depends on the polarized delivery of secretory vesicles, implying that the Myo2p tail binds to secretory vesicles. Most of the new mutations alter a region of the Myo2p tail conserved with vertebrate myosin Vs but divergent from Myo4p, the myosin V involved in mRNA transport, and genetic data suggest that the tail interacts with Smy1p, a kinesin homologue, and Sec4p, a vesicle-associated Rab protein. The data support a model in which the Myo2p tail tethers secretory vesicles, and the motor transports them down polarized actin cables to the site of exocytosis. Key words: cell polarity • myosin V • MYO2 gene product • exocytosis • Saccharomyces cerevisiae

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Abstract. To identify septin-interacting proteins in Saccharomyces cerevisiae, we screened for mutations that are synthetically lethal with a cdc12 septin mutation. One of the genes identified was GIN4, which encodes a protein kinase related to Hsl1p/Nik1p and Ycl024Wp in S. cerevisiae and to Nim1p/Cdr1p and Cdr2p in Schizosaccharomyces pombe. The Gin4p kinase domain displayed a two-hybrid interaction with the COOH-terminal portion of the Cdc3p septin, and Gin4p colocalized with the septins at the mother–bud neck. This localization depended on the septins and on the COOH-terminal (nonkinase) region of Gin4p, and overproduction of this COOH-terminal region led to a loss of septin organization and associated morphogenetic defects. We detected no effect of deleting YCL024W, either alone or in combination with deletion of GIN4. Deletion of GIN4 was not lethal but led to a striking reorganization of the septins accompanied by morphogenetic abnormalities and a defect in cell separation; however, remarkably, cytokinesis appeared to occur efficiently. Two other proteins that localize to the neck in a septin-dependent manner showed similar reorganizations and also appeared to remain largely functional. The septin organization observed in gin4� vegetative cells resembles that seen normally in cells responding to mating pheromone, and no Gin4p was detected in association with the septins in such cells. The organization of the septins observed in gin4 � cells and in cells responding to pheromone appears to support some aspects of the model for septin organization suggested previously by Field et al. (Field, C.M., O. Al-

Abstract. The conserved Ipl1 protein kinase is essential for proper chromosome segregation and thus cell viability in the budding yeast Saccharomyces cerevisiae. Its human homologue has been implicated in the tumorigenesis of diverse forms of cancer. We show here that sister chromatids that have separated from each other are not properly segregated to opposite poles of ipl1-2 cells. Failures in chromosome segregation are often associated with abnormal distribution of the spindle pole–associated Nuf2-GFP protein, thus suggesting a link between potential spindle pole defects and chromosome missegregation in ipl1 mutant cells. A small fraction of ipl1-2 cells also appears to be defective in nuclear migration or bipolar spindle formation. Ipl1 associates, probably directly, with the novel and essential

Severalproteinsequenceanalysis algorithmsarebasedonpropertiesofaminoacid compositionandrepetitiveness.Theseincludemethodsforpredictionofsecondarystructureelements, coiled-coils,transmembranesegmentsorsignalpeptides, andforassignmentoflow-complexity,nonglobular, orintrinsicallyunstructuredregions.The qualityofsuchanalysescanbegreatlyenhancedby graphicalsoftwaretoolsthatpresentpredictedsequencefeaturestogetherincontextandallowjudg - menttobefocusedsimultaneouslyonseveraldifferenttypesofsupportinginformation. Forthese purposes,wedescribetheSFINXpackage,which allowsmanydifferentsetsofsegmentalorcontinuous -curvesequencefeaturedata,generatedbyindividualexternalprograms, tobeviewedincombinationalongsideasequencedot -plotoramultiple alignmentofdatabasematches.Theimplementationiscurrentlybasedonextensionstothegraphi - calviewersDotterandBlixemandscriptsthat convertdatafromexternalprogramstoasimple genericdatadefinitionformatcalledSFS.Wedescribeapplicationsinwhichdot -plotsandflanking databasematchesprovidevaluablecontextualinformationforanalysesbasedoncompositionaland repetitivesequencefeatures.Thesystemisalso usefulforcomparingresultsfromalgorithmsrun witharangeofparameterstodetermineappropriatevaluesfordefaultsorcutoffsforlarge -scale genomicanalyses.Proteins2001;45:262--273. 2001Wiley-Liss,Inc. Keywords:sequenceanalysis;graphicalvisualization; dot-plot;databasesearchviewing; sequencecomplexity;transmembrane; coiled-coil;proteinstructure;nonglobularproteins; algorithms;datadefinition format

Abstract. The Saccharomyces cerevisiae pex17-1 mutant was isolated from a screen to identify mutants defective in peroxisome biogenesis. pex17-1 and pex17 null mutants fail to import matrix proteins into peroxisomes via both PTS1- and PTS2-dependent pathways.

REPPER (REPeats and their PERiodicities) is an integrated server that detects and analyzes regions with short gapless repeats in protein sequences or alignments. It finds periodicities by Fourier Transform (FTwin) and internal similarity analysis (REPwin). FTwin assigns numerical values to amino acids that reflect certain properties, for instance hydrophobicity, and gives information on corresponding periodicities. REPwin uses self-alignments and displays repeats that reveal significant internal similarities. Both programs use a sliding window to ensure that different periodic regions within the same protein are detected independently. FTwin and REPwin are complemented by secondary structure prediction (PSIPRED) and coiled coil prediction (COILS), making the server a versatile analysis tool for sequences of fibrous proteins. REPPER is available at

The emergence of drug-resistant strains of bacteria is a signi®cant and growing human health problem. Active ef¯ux pumps with broad speci-®city are involved in the intrinsic and acquired