Characterization and comparative analysis of Arabidopsis phosphatidylinositol phosphate 5‐kinase 10 reveals differences in Arabidopsis and human phosphatidylinositol phosphate kinases

Abstract

Arabidopsis phosphatidylinositol phosphate (PtdInsP) kinase 10 (AtPIPK10; At4g01190) is shown to be a functional enzyme of the subfamily A, type I AtPtdInsP kinases. It is biochemically distinct from AtPIPK1 (At1g21980), the only other previously characterized AtPtdInsP kinase which is of the B subfamily. AtPIPK10 has the same K m , but a 10-fold lower V max than AtPIPK1 and it is insensitive to phosphatidic acid. AtPIPK10 transcript is most abundant in inflorescence stalks and flowers, whereas AtPIPK1 transcript is present in all tissues. Comparative analysis of recombinant AtPIPK10 and AtPIPK1 with recombinant HsPIPKIa reveals that the Arabidopsis enzymes have roughly 200-and 20-fold lower V max /K m , respectively. These data reveal one explanation for the longstanding mystery of the relatively low phosphatidylinositol-(4,5)-bisphosphate:phosphatidylinositol-4-phosphate ratio in terrestrial plants.

TheArabidopsisPhosphatidylinositol Phosphate 5-Kinase PIP5K3 Is a Key Regulator of Root Hair Tip Growth

“…Recombinant PIP5K3 Protein Shows PtdIns4P 5-Kinase Activity in Vitro PIP5K3 was expected to have PtdIns4P 5-kinase activity because of its amino acid sequence similarity to PIP5K1 and PIP5K4, for which enzyme activity has been determined in vitro (Mikami et al, 1998;Elge et al, 2001;Westergren et al, 2001;Perera et al, 2005;Lee et al, 2007). To investigate the activity of PIP5K3, a fusion protein with glutathione S-transferase (GST) was expressed in E. coli cells.…”

“…Translational or posttranslational regulation may limit the PIP5K3 protein to developing root hair cells. For the other PIP5K genes, such a specific expression pattern has not been observed (Mikami et al, 1998;Perera et al, 2005;Lee et al, 2007;Lou et al, 2007), suggesting that the PIP5K3 gene has evolved to have a specific function in root hair development. To date, the biological functions of PIP5K1 and PIP5K9 have been assumed to be signal transduction for overall cell responses to abscisic acid and sugar, respectively (Mikami et al, 1998;Elge et al, 2001;Lou et al, 2007).…”

“…Of the 11 PIP5Ks in Arabidopsis, only the in vitro enzyme activity of PIP5K1, PIP5K4, and PIP5K10 have been characterized, preferentially catalyzing PtdIns(4,5)P 2 formation from PtdIns4P (Mikami et al, 1998;Elge et al, 2001;Westergren et al, 2001;Perera et al, 2005;Lee et al, 2007). The GST-PIP5K3 fusion protein also showed the 5-kinase activity in vitro, with a substrate preference for PtdIns4P.…”

A Dibasic Amino Acid Pair Conserved in the Activation Loop Directs Plasma Membrane Localization and Is Necessary for Activity of Plant Type I/II Phosphatidylinositol Phosphate Kinase

“…Although PA stimulation of the animal type I and plant type I/II PIPKs has been shown (Moritz et al, 1992;Jenkins et al, 1994;Ishihara et al, 1998;Perera et al, 2005;Im et al, 2007;Saavedra et al, 2009), the mechanism by which PA stimulates PIPK activity has yet to be studied in detail. However, Stace et al (2008) recently demonstrated that basic amino acids upstream of the activation loop in the catalytic region are involved in PA-dependent activation of human PIP5K-1b.…”

“…Since the corresponding amino acid residue is Glu in all plant PIPKs so far reported, it is suggested that there also is a plantspecific mode of substrate specificity regulation in plant type I/II PIPKs. However, enzymatic activity appears to be modified in similar ways between plant type I/II and animal type I PIPKs; that is, phosphorylation-and PA-dependent activation of PIPKs has been observed in both animals and plants (Moritz et al, 1992;Jenkins et al, 1994;Pical et al, 1999;Westergren et al, 2001;Perera et al, 2005;Saavedra et al, 2009).…”