Phosphatidylinositol 4-kinase: Gene structure and requirement for yeast cell viability

Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Science (Impact Factor: 33.61). 12/1993; 262(5138):1444-8. DOI: 10.1126/science.8248783
Source: PubMed


Phosphatidylinositol (PtdIns) 4-kinase catalyzes the first step in the biosynthesis of PtdIns-4,5-bisphosphate (PtdIns[4,5]P2).
Hydrolysis of PtdIns[4,5]P2 in response to extracellular stimuli is thought to initiate intracellular signaling cascades that
modulate cell proliferation and differentiation. The PIK1 gene encoding a PtdIns 4-kinase from the yeast Saccharomyces cerevisiae
was isolated by polymerase chain reaction (PCR) with oligonucleotides based on the sequence of peptides derived from the purified
enzyme. The sequence of the PIK1 gene product bears similarities to that of PtdIns 3-kinases from mammals (p110) and yeast
(Vps34p). Expression of PIK1 from a multicopy plasmid elevated PtdIns 4-kinase activity and enhanced the response to mating
pheromone. A pik1 null mutant was inviable, indicating that PtdIns4P and presumably PtdIns[4,5]P2 are indispensable phospholipids.

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    • "The inositol used in PI synthesis is either synthesized de novo (discussed below) or obtained from the growth medium via the ITR1-and ITR2-encoded inositol transporters (Table 2) (Nikawa et al. 1991). Once formed, PI may be converted to PI 3-P by the VPS34-encoded PI 3 kinase (Herman and Emr 1990; Schu et al. 1993) or to PI 4-P by the PI 4 kinases encoded by LSB6 (Han G-S et al. 2002; Shelton et al. 2003), STT4 (Yoshida et al. 1994a), and PIK1 (Flanagan et al. 1993; Garcia-Bustos et al. 1994). PI 4-P may be further phosphorylated to PI 4,5-P 2 by the MSS4- encoded PI 4-P 5 kinase (Yoshida et al. 1994b), whereas PI 3-P may be phosphorylated to PI 3,5-P 2 by the FAB1- encoded PI 3-P 5 kinase (Yamamoto et al. 1995). "
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    ABSTRACT: Due to its genetic tractability and increasing wealth of accessible data, the yeast Saccharomyces cerevisiae is a model system of choice for the study of the genetics, biochemistry, and cell biology of eukaryotic lipid metabolism. Glycerolipids (e.g., phospholipids and triacylglycerol) and their precursors are synthesized and metabolized by enzymes associated with the cytosol and membranous organelles, including endoplasmic reticulum, mitochondria, and lipid droplets. Genetic and biochemical analyses have revealed that glycerolipids play important roles in cell signaling, membrane trafficking, and anchoring of membrane proteins in addition to membrane structure. The expression of glycerolipid enzymes is controlled by a variety of conditions including growth stage and nutrient availability. Much of this regulation occurs at the transcriptional level and involves the Ino2-Ino4 activation complex and the Opi1 repressor, which interacts with Ino2 to attenuate transcriptional activation of UAS(INO)-containing glycerolipid biosynthetic genes. Cellular levels of phosphatidic acid, precursor to all membrane phospholipids and the storage lipid triacylglycerol, regulates transcription of UAS(INO)-containing genes by tethering Opi1 to the nuclear/endoplasmic reticulum membrane and controlling its translocation into the nucleus, a mechanism largely controlled by inositol availability. The transcriptional activator Zap1 controls the expression of some phospholipid synthesis genes in response to zinc availability. Regulatory mechanisms also include control of catalytic activity of glycerolipid enzymes by water-soluble precursors, products and lipids, and covalent modification of phosphorylation, while in vivo function of some enzymes is governed by their subcellular location. Genome-wide genetic analysis indicates coordinate regulation between glycerolipid metabolism and a broad spectrum of metabolic pathways.
    Genetics 02/2012; 190(2):317-49. DOI:10.1534/genetics.111.130286 · 5.96 Impact Factor
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    • "This nonconservative missense change substitutes a basic amino acid for an acidic residue. Furthermore, Glu 725 is a highly conserved residue in the PIK domain of the AGE-1 protein, which is a homolog of the vertebrate PI3 kinase (Flanagan et al. 1993; Morris et al. 1996; Domin and Waterfield 1997) (Figure 3). This mutation segregated with the lifespan extension phenotype after four rounds of backcrossing (data not shown), supporting the conclusion that the missense change resulting in the E725K substitution is the am88 mutation, and am88 is a novel allele of age-1. "
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    ABSTRACT: Aging is an important feature of animal biology characterized by progressive, degenerative changes in somatic and reproductive tissues. The rate of age-related degeneration is genetically controlled, since genes that influence lifespan have been identified. However, little is known about genes that affect reproductive aging or aging of specific somatic tissues. To identify genes that are important for controlling these degenerative changes, we used chemical mutagenesis to perform forward genetic screens in Caenorhabditis elegans. By conducting a screen focused on somatic aging, we identified mutant hermaphrodites that displayed extended periods of pharyngeal pumping, body movement, or survival. One of these mutations is a novel allele of the age-1 gene. age-1 encodes a phosphatidylinositol-3-kinase (PI3K) that functions in the insulin/insulin-like growth factor-1 (IGF-1) signaling pathway. age-1(am88) creates a missense change in the conserved PIK domain and causes dramatic extensions of the pharyngeal pumping and body movement spans, as well as a twofold extension of the lifespan. By conducting screens focused on reproductive aging in mated hermaphrodites, we identified mutants that displayed increased progeny production late in life. To characterize these mutations, we developed quantitative measurements of age-related morphological changes in the gonad. The am117 mutation delayed age-related declines in progeny production and morphological changes in the gonad. These studies provide new insights into the genetic regulation of age-related degenerative changes in somatic and reproductive tissues.
    Genetics 07/2011; 189(1):341-56. DOI:10.1534/genetics.111.130450 · 5.96 Impact Factor
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    • "The type-III PI4Ks are relatives of the PI 3-kinase family, while the smaller type-II enzymes form a separate family [12] [13]. The first cloned PI4Ks were the yeast orthologues of the type-III PI4Ks [14]. PIK1 was recognized as an essential gene important in Golgi to plasma membrane secretion [15] [16] whereas another yeast PI4K, STT4 was discovered as a gene whose mutations cause staurosporine hypersensitivity [17]. "
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    ABSTRACT: Phosphatidylinositol 4-kinase type IIIa (PI4KIIIα) is one of four mammalian PI 4-kinases that catalyzes the first committed step in polyphosphoinositide synthesis. PI4KIIIα has been linked to regulation of ER exit sites and to the synthesis of plasma membrane phosphoinositides and recent studies have also revealed its importance in replication of the Hepatitis C virus in liver. Two isoforms of the mammalian PI4KIIIα have been described and annotated in GenBank: a larger, ~230kDa (isoform 2) and a shorter splice variant containing only the ~97kDa C-terminus that includes the catalytic domain (isoform 1). However, Northern analysis of human tissues and cancer cells showed only a single transcript of ~7.5kb with the exception of the proerythroleukemia line K562, which contained significantly higher level of the 7.5kb transcript along with smaller ones of 2.4, 3.5 and 4.2kb size. Bioinformatic analysis also confirmed the high copy number of PI4KIIIα transcript in K562 cells along with several genes located in the same region in Chr22, including two pseudogenes that cover most exons coding for isoform 1, consistent with chromosome amplification. A panel of polyclonal antibodies raised against peptides within the C-terminal half of PI4KIIIα failed to detect the shorter isoform 1 either in COS-7 cells or K562 cells. Moreover, expression of a cDNA encoding isoform 1 yielded a protein of ~97kDa that showed no catalytic activity and failed to rescue hepatitis C virus replication. These data draw attention to PI4KIIIα as one of the genes found in Chr22q11, a region affected by chromosomal instability, but do not substantiate the existence of a functionally relevant short form of PI4KIIIα.
    Biochimica et Biophysica Acta 07/2011; 1811(7-8):476-83. DOI:10.1016/j.bbalip.2011.04.013 · 4.66 Impact Factor
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