1887

Abstract

The early endocytic patch protein Sla2 is important for morphogenesis and growth rates in and but the mechanism that connects these processes is not clear. Here we report that growth defects in cells lacking Ca or Sc are associated with a cell cycle delay that is influenced by Swe1, a morphogenesis checkpoint kinase. To establish how Swe1 monitors Sla2 function, we compared actin organization and cell cycle dynamics in strains lacking other components of early endocytic patches (Sla1 and Abp1) with those in strains lacking Sla2. Only strains had defects in actin cables, a known trigger of the morphogenesis checkpoint, yet all three strains exhibited Swe1-dependent phenotypes. Thus, Swe1 appears to monitor actin patch in addition to actin cable function. Furthermore, Swe1 contributed to virulence in a mouse model of disseminated candidiasis, implying a role for the morphogenesis checkpoint during the pathogenesis of infections.

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2009-12-01
2019-10-20
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[ ] [ ] (.mov files, 369 and 136 kb, respectively). Representative fluorescence time-lapse images of C. albicans wild-type (YJB6709) and sla2 (YJB6757) mutant strains expressing Abp1-YFP to mark actin patches. Individual images were collected at 1 s intervals. Fluorescent protein fusions were constructed by PCR-mediated gene modification using ABP1-specific primers as described in Methods. Quantification of actin patch dynamics in strains described in the legend to movies 1 and 2. 'Duration' is the average time that a given patch was visible. 'Distance' is the total distance that patches moved during their existence. 'Rate' is the average distance a patch moved per sec, discarding frames where a patch had no movement. * ≤0.001 by Student's -test. [ PDF] (29 kb) Sla2 colocalizes with a subset of actin patches. YJB6709 expressing Sla2-YFP was grown to exponential phase and stained with rhodamine-phalloidin as described in Methods. Representative DIC and fluorescence images are shown. The right-most panel shows the merged image from the YFP and rhodamine-phalloidin panels, with the yellow signal indicating colocalization. [ PDF] (197 kb)

MOVIE

[ ] [ ] (.mov files, 369 and 136 kb, respectively). Representative fluorescence time-lapse images of C. albicans wild-type (YJB6709) and sla2 (YJB6757) mutant strains expressing Abp1-YFP to mark actin patches. Individual images were collected at 1 s intervals. Fluorescent protein fusions were constructed by PCR-mediated gene modification using ABP1-specific primers as described in Methods. Quantification of actin patch dynamics in strains described in the legend to movies 1 and 2. 'Duration' is the average time that a given patch was visible. 'Distance' is the total distance that patches moved during their existence. 'Rate' is the average distance a patch moved per sec, discarding frames where a patch had no movement. * ≤0.001 by Student's -test. [ PDF] (29 kb) Sla2 colocalizes with a subset of actin patches. YJB6709 expressing Sla2-YFP was grown to exponential phase and stained with rhodamine-phalloidin as described in Methods. Representative DIC and fluorescence images are shown. The right-most panel shows the merged image from the YFP and rhodamine-phalloidin panels, with the yellow signal indicating colocalization. [ PDF] (197 kb)

MOVIE

[ ] [ ] (.mov files, 369 and 136 kb, respectively). Representative fluorescence time-lapse images of C. albicans wild-type (YJB6709) and sla2 (YJB6757) mutant strains expressing Abp1-YFP to mark actin patches. Individual images were collected at 1 s intervals. Fluorescent protein fusions were constructed by PCR-mediated gene modification using ABP1-specific primers as described in Methods. Quantification of actin patch dynamics in strains described in the legend to movies 1 and 2. 'Duration' is the average time that a given patch was visible. 'Distance' is the total distance that patches moved during their existence. 'Rate' is the average distance a patch moved per sec, discarding frames where a patch had no movement. * ≤0.001 by Student's -test. [ PDF] (29 kb) Sla2 colocalizes with a subset of actin patches. YJB6709 expressing Sla2-YFP was grown to exponential phase and stained with rhodamine-phalloidin as described in Methods. Representative DIC and fluorescence images are shown. The right-most panel shows the merged image from the YFP and rhodamine-phalloidin panels, with the yellow signal indicating colocalization. [ PDF] (197 kb)

PDF

[ ] [ ] (.mov files, 369 and 136 kb, respectively). Representative fluorescence time-lapse images of C. albicans wild-type (YJB6709) and sla2 (YJB6757) mutant strains expressing Abp1-YFP to mark actin patches. Individual images were collected at 1 s intervals. Fluorescent protein fusions were constructed by PCR-mediated gene modification using ABP1-specific primers as described in Methods. Quantification of actin patch dynamics in strains described in the legend to movies 1 and 2. 'Duration' is the average time that a given patch was visible. 'Distance' is the total distance that patches moved during their existence. 'Rate' is the average distance a patch moved per sec, discarding frames where a patch had no movement. * ≤0.001 by Student's -test. [ PDF] (29 kb) Sla2 colocalizes with a subset of actin patches. YJB6709 expressing Sla2-YFP was grown to exponential phase and stained with rhodamine-phalloidin as described in Methods. Representative DIC and fluorescence images are shown. The right-most panel shows the merged image from the YFP and rhodamine-phalloidin panels, with the yellow signal indicating colocalization. [ PDF] (197 kb)

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