Activities and Structure-Function Analysis of Fission Yeast Inositol Pyrophosphate (IPP) Kinase-Pyrophosphatase Asp1 and Its Impact on Regulation of pho1 Gene Expression

Abstract

Expression of the fission yeast phosphate regulon is sensitive to the intracellular level of the inositol pyrophosphate (IPP) signaling molecule 1,5-IP 8 . IP 8 dynamics are determined by Asp1, a bifunctional enzyme comprising N-terminal IPP 1-kinase and C-terminal IPP 1-pyrophosphatase domains that catalyze IP 8 synthesis and catabolism, respectively.

Inositol pyrophosphate-controlled kinetochore architecture and mitotic entry in S. pombe

“…We and others have described the catalytic properties of recombinant Asp1 purified from bacterial expression systems [7, 10, 31, 36, 74]. In these in vitro assays, Asp1 kinase and pyrophosphatase domains can independently interact with IPP substrates and perform their catalytic functions at position C1 of the inositol ring before releasing their products (shown diagrammatically in Figure 1A).…”

“…As other members of the Vip1/PPIP5K family, the fission yeast Schizosaccharomyces pombe Asp1 protein is a bifunctional enzyme consisting of an N-terminal kinase domain with 1-kinase activity generating the less abundant IPP IP8 and a [2Fe-2S]-binding C-terminal pyrophosphatase domain with specific inositol pyrophosphate 1-phosphatase activity (Figure 1A [7,10,13,[35][36][37]. Asp1 was originally discovered as a modulator of the cortical actin cytoskeleton, but since then numerous biological functions have been identified to be regulated by this protein [6,7,13,19,[38][39][40][41].…”

“…The functions of these molecules are very diverse, ranging from defense against pathogens in humans and plants, to mammalian organ development and fungal morphogenesis and pathogenicity [2][3][4][5][6][7][8]. Additionally, IPPs play an important function in cell adaptation to adverse environmental conditions, including a predominant role in maintaining phosphate homeostasis in mammals, plants and fungi [4,[9][10][11][12][13][14][15][16][17][18][19]. Although, IPPs appear to have rather pleiotropic effects, evidence is accumulating that different phenotypes are connected such as fungal virulence and phosphate homeostasis [20].…”

Structures of Fission Yeast Inositol Pyrophosphate Kinase Asp1 in Ligand-Free, Substrate-Bound, and Product-Bound States

“…IP 8 is generated from phytic acid (IP 6 ) by the sequential action of fission yeast kinases Kcs1, which converts IP 6 to 5-IP 7 , and Asp1, which converts 5-IP 7 to 1,5-IP 8 ( 4 , 5 ). The PHO genes are dysregulated by genetic manipulations of Asp1, a bifunctional kinase and pyrophosphatase enzyme that consists of an N-terminal kinase domain that synthesizes 1,5-IP 8 and a C-terminal pyrophosphatase domain that converts IP 8 back to 5-IP 7 ( 6 – 8 ). A pyrophosphatase-defective asp1-H397A allele (with a His-to-Ala change at residue 397) that increases the intracellular level of IP 8 ( 6 , 7 ) derepresses pho1 expression by favoring precocious 3′-processing and termination of flanking prt lncRNA synthesis in response to poly(A) signals upstream of the mRNA promoter, in a manner dependent on the cleavage and polyadenylation factor (CPF) complex and transcription termination factor Rhn1 ( 1 ).…”

“…The isolated N-terminal kinase domains of Asp1, Vip1, PPIP5K, and VIH have autonomous kinase activity ( 6 – 8 , 13 – 15 ). Wang et al conducted incisive structural and mechanistic studies of the kinase domain of human PPIP5K2 ( 16 ).…”