Identification of protein phosphatases dephosphorylating mRNP proteins from cryptobiotic gastrulae of the brine shrimp A.salina

Thoen, Chris; Hove, Lauris Van; Cohen, Philip; Slegers, Herman

doi:10.1016/0006-291x(85)91773-5

Cited by 8 publications

(2 citation statements)

References 23 publications

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“…In the bIine shrimp, PP-2A dephosphorylates messenger ribonucleoproteins phosphorylated by a casein kinase-II activity (32,33), another level at which protein synthesis could be controlled.…”

Section: Muscle Contractilitymentioning

confidence: 99%

The Structure and Regulation of Protein Phosphatases

Cohen

1989

Annu. Rev. Biochem.

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Four major serine/threonine-specific protein phosphatase catalytic subunits are present in the cytoplasm of animal cells. Three of these enzymes, PP-1, PP-2A, and PP-2B, are members of the same gene family, while PP-2C appears to be distinct. PP-1, PP-2A, and PP-2B are complexed to other subunits in vivo, whereas PP-2C has only been isolated as a monomeric protein. PP-1, PP-2A, and PP-2C have broad and overlapping specificities in vitro, and account for virtually all measurable activity in tissue extracts toward a variety of phosphoproteins that regulate metabolism, muscle contractility, and other processes. Their precise functions in vivo are unknown, although important clues to the physiological roles of PP-1 and PP-2A are provided by the effects of okadaic acid and by the subcellular localization of PP-1. The active forms of PP-1 are largely particulate, and their association with subcellular structures is mediated by "targetting subunits" that direct PP-1 to particular locations, enhance its activity toward certain substrates, and confer important regulatory properties upon it. This concept is best established for the glycogen-bound enzymes in skeletal muscle and liver (PP-1G) and the myofibrillar form (PP-1M) in skeletal muscle. The activities of PP-1 and PP-2B are controlled by the second messengers cyclic AMP and calcium. The activity of PP-2B is dependent on calcium and calmodulin, while PP-1 is controlled in a variety of ways that depend on the form of the enzyme and the tissue. PP-1 can be inhibited by cyclic AMP in a variety of cells through the A-kinase-catalyzed phosphorylation of inhibitor-1 and its isoforms. Phosphorylation of the glycogen-binding subunit of PP-1G by A-kinase promotes translocation of the catalytic subunit from glycogen particles to cytosol in skeletal muscle, inhibiting the dephosphorylation of glycogen-metabolizing enzymes. Allosteric inhibition of hepatic PP-1G by phosphorylase a occurs in response to signals that elevate cyclic AMP or calcium, and prevents the activation of glycogen synthase in liver. PP-1 can also be activated indirectly by calcium through the ability of PP-2B to dephosphorylate inhibitor-1. This control mechanism may operate in dopaminoceptive neurones of the brain and other cells. The inactive cytosolic form of PP-1 (PP-1I) can be activated in vitro through the glycogen synthase kinase-3-catalyzed phosphorylation of its inhibitory subunit (inhibitor-2), but the physiological significance is unclear.(ABSTRACT TRUNCATED AT 400 WORDS)

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Section: Muscle Contractilitymentioning

confidence: 99%

The Structure and Regulation of Protein Phosphatases

Cohen

1989

Annu. Rev. Biochem.

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2,484

1,651

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“…In an attempt to characterize the catalytic subunit of the protein phosphatase, the enzyme (after step 10) was precipitated with 80% (v/v) ethanol at room temperature [37]. This treatment results in a threefold increase in the rate of dephosphorylation [41]. When the ethanol-treated enzyme was chromatographed on Spherogel TSK G3000 SW, the phosphatase activity emerged as one symmetrical peak, which probably corresponds to the catalytic p subunit with an M, of 35000 & 4000 (Fig.…”

Section: Purity Molecular Muss and Subunit Structure Of The Enzymementioning

confidence: 99%

Purification, subunit structure and properties of a high-molecular-mass protein phosphatase capable of dephosphorylating mRNP of the brine shrimp Artemia sp.

1987

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A phosphoprotein phosphatase active towards casein, phosphorylase a and mRNP proteins has been detected in the cytosol of cryptobiotic gastrulae of Artemia sp. This phosphatase has a relative molecular mass ( M , ) of 225000 as measured by gel filtration on Sephadex G-200 and has been purified to near homogeneity by ionexchange chromatography on different DEAE-substituted matrices, affinity chromatography on polylysineagarose, histone-Sepharose 4B and protamine-agarose, hydrophobic chromatography on phenyl-Sepharose 4B and gel filtration on Sephadex G-200. Sodium dodecyl sulphate gel electrophoresis of the final purification step revealed that the enzyme contains two types of subunits, CI and jl, with M , of 40000 and 75000, respectively. These values, in conjunction with the native M , and the molar ratios of the subunits estimated by densitometric analysis of the gel, suggested that the subunit composition of the enzyme is a2fi2. When treated with 1.7% (v/v) 2-mercaptoethanol at -20°C or with ethanol, the enzyme released the catalytic a subunit of M , 40000. = 1.6 pM) and polylysine (Ao.s = 0.2 pM) and inhibited by ATP (10,5 = 12 pM), NaF = 3.1 mM) and pyrophosphate (fo.5 = 0.6 mM). The enzyme is a polycation-stimulated protein phosphatase. Purified mRNP proteins, phosphorylated by the mRNP-associated casein kinase type 11, are among the substrates used by the enzyme. The function of reversible phosphorylation-dephosphorylation of mRNP as a regulatory mechanism in mRNP metabolism is discussed.The protein phosphatase was activated by basic proteins e.g. protamine (Ao.5 = 1 pM), histone H I Protein phosphorylation-dephosphorylation has emerged as a regulatory mechanism modifying the action of enzymes and structural proteins [I]. Rate-limiting enzymes in biodegradative pathways are usually activated by phosphorylation, whereas those in biosynthetic pathways are generally inactivated [2, 31. There is increasing evidence to support the hypothesis that different metabolic events are regulated by relatively few protein kinases and phosphatases [l, phorylate the a subunit of phosphorylase kinase and are insensitive to inhibitors-1 and -2 (protein phosphatases-2A, 2B and 2C) [17,18]. Based on enzyme-directed regulation, protein phosphatases may also be classified into four main classes: ATP/Mg-dependent protein phosphatases, calcineurin, Mg2+-dependent protein phosphatases and polycationstimulated protein phosphatases [19].Messenger RNA is associated with specific proteins to form messenger ribonucleoproteins (mRNP) [20]. It is generally accepted that the mRNA-associated proteins have a function in mRNA metabolism, e. g. replication, stability, storage, transport and translation [21-251. Phosphorylation of mRNP-associated proteins has been observed frequently and is proposed to regulate mRNA metabolism [26 -291. Furthermore, Egly and coworkers [27,30] have suggested that dephosphorylation of mRNP proteins also has a function in translation.Cryptobiotic gastrulae, a dormant phase of the embryos of the brine shrimp ...

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Identification of the substrates of the casein kinase II associated with non-polysomal messenger ribonucleoproteins of A. salina cryptobiotic embryos

Thoen¹,

Hove²,

Slegers³

1986

Molec. Biol. Rep.

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The association of a protein kinase with cytoplasmic non-polysomal messenger ribonucleoproteins is demonstrated by chromatography on oligo(dT)-cellulose and sucrose gradient centrifugation. The cAMP-independent enzyme is inhibited by caffeine and poly(L)-glutamic acid and is classified as a casein kinase II. Among the exogenous proteins initiation factor eIF2 is the best substrate and is 7.8 times more efficiently phosphorylated than casein. Endogenous mRNP protein substrates have a Mr of 125 000, 65 000, 38 000, 26 000 and 23 500. The main phosphate acceptor is the Mr 38 000 poly(A)-binding protein. Dephosphorylation of the poly(A)-binding protein by protein phosphatases decreases its RNA binding property. The effect of phosphorylation-dephosphorylation of mRNP proteins on the initiation of protein synthesis is discussed.

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Identification of protein phosphatases dephosphorylating mRNP proteins from cryptobiotic gastrulae of the brine shrimp A.salina

Cited by 8 publications

References 23 publications

The Structure and Regulation of Protein Phosphatases

The Structure and Regulation of Protein Phosphatases

Purification, subunit structure and properties of a high-molecular-mass protein phosphatase capable of dephosphorylating mRNP of the brine shrimp Artemia sp.

Identification of the substrates of the casein kinase II associated with non-polysomal messenger ribonucleoproteins of A. salina cryptobiotic embryos

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