An SH2-domain-containing kinase negatively regulates the phosphatidylinositol-3 kinase pathway.
SHK1 is a novel dual-specificity kinase that contains an SH2 domain in its C-terminal region. We demonstrate that SHK1 is required for proper chemotaxis and phagocytosis. Mutant shk1 null cells lack polarity, move very slowly, and exhibit an elevated and temporally extended chemoattractant-mediated activation of the kinase Akt/PKB. GFP fusions of the PH domain of Akt/PKB or the PH-domain-containing protein CRAC, which become transiently associated with the plasma membrane after a global stimulation with a chemoattractant, remain associated with the plasma membrane for an extended period of time in shk1 null cells. These results suggest that SHK1 is a negative regulator of the PI3K (phosphatidylinositol-3 kinase) pathway. Furthermore, when a chemoattractant gradient is applied to a wild-type cell, these PH-domain-containing proteins and the F-actin-binding protein coronin localize to its leading edge, but in an shk1 null cell they become randomly associated with the plasma membrane and cortex, irrespective of the direction of the chemoattractant gradient, suggesting that SHK1 is required for the proper spatiotemporal control of F-actin levels in chemotaxing cells. Consistent with such functions, SHK1 is localized at the plasma membrane/cortex, and we show that its SH2 domain is required for this localization and the proper function of SHK1.
Section of Cell and Developmental Biology and Center for Molecular Genetics, University of California, San Diego, La Jolla, California 92093-0634, USA.
J
J
Moniakis
S
S
Funamoto
M
M
Fukuzawa
J
J
Meisenhelder
T
T
Araki
T
T
Abe
R
R
Meili
T
T
Hunter
J
J
Williams
R A
RA
Firtel
D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002633:Chemotaxis / Q000235:genetics; D000242:Cyclic AMP / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D004023:Dictyostelium / Q000378:metabolism; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D008840:Microfilament Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D010587:Phagocytosis; D010590:Phalloidine / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D019606:Phosphoamino Acids / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D019076:Transgenes; D054462:p21-Activated Kinases; D018909:src Homology Domains
D000199:Actins; D000242:Cyclic AMP; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D008840:Microfilament Proteins; D010590:Phalloidine; D019869:Phosphatidylinositol 3-Kinases; D019606:Phosphoamino Acids; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011993:Recombinant Fusion Proteins; D029702:Schizosaccharomyces pombe Proteins; D054462:p21-Activated Kinases; C072061:coronin proteins; C094341:shk1 protein, S pombe