Saccharomyces cerevisiae contains a Type II phosphoinositide 4-kinase

Department of Pharmacology, University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Blvd., Dallas, TX 75390-9041, USA.
Biochemical Journal (Impact Factor: 4.4). 05/2003; 371(Pt 2):533-40. DOI: 10.1042/BJ20021407
Source: PubMed


The yeast Saccharomyces cerevisiae contains two known phosphoinositide 4-kinases (PI 4-kinases), which are encoded by PIK1 and STT4; both are essential. Pik1p is important for exocytic transport from the Golgi, whereas Stt4p plays a role in cell-wall integrity and cytoskeletal rearrangements. In the present study, we report that cells have a third PI 4-kinase activity encoded by LSB6, a protein identified previously in a two-hybrid screen as interacting with LAS17p. Although Pik1p and Stt4p are closely related members of the Type III class of PI 4-kinases, Lsb6p belongs to the distinct Type II class, based on its amino acid sequence, its sensitivity to inhibition by adenosine and its insensitivity to wortmannin. Lsb6p is the first fungal Type II enzyme cloned. The protein was expressed and purified from Sf9 cells and used to define kinetic parameters. As commonly observed for surface-active enzymes, activities varied both with substrate concentration and lipid/detergent molar ratios. Maximal activities of approx. 100 min(-1) were obtained at the PI/Triton X-100 ratio of 1:5. The K (m) value for ATP was 266 microM, intermediate between the values reported for mammalian Type II and III kinases. Epitope-tagged protein, expressed in yeast, was entirely particulate, and about half of it could be extracted with non-ionic detergent. Lsb6p-green fluorescent protein was found both on vacuolar membranes and on the plasma membrane, suggesting a role in endocytic or exocytic pathways.

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Available from: Joel M Goodman, Feb 24, 2014
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    • "Phosphoinositide kinases, such as PI 4-kinases or PI4P 5-kinases, can each be represented by different subfamilies of enzymes, that can have non-redundant functionality. For instance, in the yeast Saccharomyces cerevisiae three PI 4-kinases with non-redundant functions exist (Table 1) which represent two distinct enzyme classes (Shelton et al. 2003; Demmel et al. 2008). In A. thaliana, four ubiquitously expressed homologs of the yeast enzymes "
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    • "The third yeast PI4K, Lsp6p, also termed Pik2p, is a g-group member. Loss of Lsp6p is not lethal and leads to only moderate alterations in PtdIns4P levels (Han et al. 2002; Shelton et al. 2003). Two mammalian a-group members are isoforms of a single gene, PI4KIIIa, with isoform 1 containing just the c-terminal portion of the protein. "
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    • "Before our study, Lsb6, the sole type II PI 4-kinase of yeast, had been characterized biochemically (Han et al., 2002; Shelton et al., 2003). However, its function was unknown because lsb6⌬ mutants did not exhibit growth defects (Han et al., 2002; Shelton et al., 2003), in contrast to the lethal growth deficits of mutants defective in the type III PI 4-kinases Stt4 or Pik1. However, we show herein that Lsb6 is required for the motility of endosomes containing Ste2, a novel function for type II PI 4-kinases. "
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