Targeted Disruption of the Mouse PAS Domain Serine/Threonine Kinase PASKIN

Abstract

The PAS (Per-Arnt-Sim) domain is a widespread protein fold of environmental protein sensors involved in the perception of light intensity, oxygen partial pressure, redox potentials, and voltage (9,18,26,29). While most PAS proteins are known from the archaea, bacteria, and lower eukarya, relatively few sensor PAS proteins have been reported in mammalian species. Apart from the HERG voltage-dependent potassium channel (15), the mammalian PAS domain-containing proteins identified so far serve as heterodimerization interfaces of transcription factors involved in the xenobiotic response (25), adaptation to hypoxia (28), circadian rhythm generation (19), and possibly, carbon monoxide signaling (7).By database searches with the PAS sequence as a bait, we and others previously identified a novel mammalian PAS protein, termed PASKIN (10) or PAS kinase (22). The domain architecture of PASKIN resembles that of the oxygen sensor protein FixL from Rhizobium species which contains a hemebearing PAS domain and a histidine kinase domain that couples sensing to signaling (8). PASKIN contains two PAS domains, with higher sequence similarity to the FixL PAS domain than to any other known PAS domain, and a serine/threonine kinase domain related to AMP kinases. As known from FixL, the PAS A domain of PASKIN represses the kinase activity in cis. Following derepression, presumably by ligand-binding to the PAS domain, autophosphorylation in trans results in the switching on of the kinase domain of PASKIN (22).Of note, the human PASKIN and mouse Paskin genes share a relatively small promoter region with PPP1R7 and Ppp1r7, respectively, suggesting coexpression of the two genes (10). PPP1R7/Ppp1r7 encodes for Sds22, the regulatory subunit 7 involved in target protein recognition of the serine/threonine protein phosphatase 1 (PP1) (2-4). Thus, it is tempting to speculate that Sds22 directs PP1-mediated dephosphorylation of activated PASKIN, resulting in the switching off of the PASKIN kinase activity.The three-dimensional structure of the PASKIN PAS A domain has recently been resolved, and synthetic ligands binding to this domain were identified (1). Ligand binding as well as mutation of the PAS A domain result in the activation of the kinase domain. The synthetic ligands identified are structurally related to dioxin, known to bind to the PAS domain of the dioxin receptor. However, an endogenous ligand of PASKIN has not been identified so far. While the physiological function of mammalian PASKIN is unknown, important insights into the function of the two Saccharomyces cerevisiae homologs have recently been obtained (23). The yeast PASKIN homologs phosphorylate three translation factors and two enzymes involved in the regulation of glycogen and trehalose synthesis, thereby coordinately controlling translation and

Involvement of Per–Arnt–Sim (PAS) kinase in the stimulation of preproinsulin and pancreatic duodenum homeobox 1 gene expression by glucose

“…mRNA encoding PASK is expressed in most mouse and human tissues (3), with particularly high levels in the thymus and testis (7). Although mice inactivated for PASK expression are normally fertile and display no apparent defects in growth and development (7), the impact of PASK deletion on glucose homeostasis or adaptation to metabolic stress is unknown.…”

PAS kinase is required for normal cellular energy balance

“…PASK Ϫ/Ϫ mice were genotyped as described (17). After the fifth backcross into C57BL/6 (Charles River Laboratories, Wilmington, MA), 12-to 24-week-old males were used for all experiments except that the islet perifusion studies were performed on females.…”

PAS Kinase Is a Nutrient and Energy Sensor in Hypothalamic Areas Required for the Normal Function of AMPK and mTOR/S6K1

“…The animals used were 10-to 16-week-old males (25-30 g), C57Bl/6 wild-type and PASK-deficient mice backcrossed into C57Bl/6 for at least 12 generations [6]. The animals were fed ad libitum with a standard pellet diet and housed at a constant temperature (21°C) on a 12-h light-dark cycle, with lights on at 8 a.m.…”

“…Hypothalamic slices were fixed in 0.2 % glutaraldehyde, 5 mM EGTA, 2 mM MgCl 2 and 0.1 M Na-phosphate buffer (pH 7.3) for 16 h. Then, they were permeabilized in 0.01 % Na-deoxycholate, 0.02 % NP-40, 2 mM MgCl 2 and 0.1 M Na-phosphate buffer (pH 7.3) and incubated in X-Gal solution [5 mM K 3 Fe(CN) 6 , 5 mM K 4 Fe(CN) 6 , 2 mM MgCl 2 , 0.02 % NP-40, 0.01 % Nadeoxycholate, 0.1 M Na-phosphate buffer (pH 7.3), 0.1 % XGal] for at least 5 h at 30°C until colour developed.…”

“…PASK could regulate glucose metabolism and cellular energy homeostasis [4,5]. Initially, PASK-deficient mice showed no developmental, growth, or reproductive defects [6]. Later investigation indicated that PASK-deficient mice are resistant to dietinduced obesity, and PASK knockdown by RNAi in cultured myoblasts causes increased glucose and palmitate oxidation and elevated cellular ATP levels [7].…”