%0 Journal Article %1 Humrich2003 %A Humrich, J. %A Bermel, C. %A Grubel, T. %A Quitterer, U. %A Lohse, M. J. %D 2003 %J J Biol Chem %K & Alternative Animals Carrier Casein Cell Enzyme Humans II Inhibitors/pharmacology Kinase Kinases/antagonists Line Mice Nerve Organ Phosphorylation/drug Protein-Serine-Threonine Proteins/genetics/*metabolism Specificity Splicing Substrate Tissue effects inhibitors/genetics/*metabolism %N 7 %P 4474-81 %T Regulation of phosducin-like protein by casein kinase 2 and N-terminal splicing %U http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12466282 %V 278 %X Phosducin-like protein (PhLP) is a member of the phosducin family of G-protein betagamma-regulators and exists in two splice variants. The long isoform PhLP(L) and the short isoform PhLP(S) differ by the presence or absence of an 83-amino acid N terminus. In isolated biochemical assay systems, PhLP(L) is the more potent Gbetagamma-inhibitor, whereas the functional role of PhLP(S) is still unclear. We now report that in intact HEK 293 cells, PhLP(S) inhibited Gbetagamma-induced inositol phosphate generation with approximately 20-fold greater potency than PhLP(L). Radiolabeling of transfected HEK 293 cells with (32)P revealed that PhLP(L) is constitutively phosphorylated, whereas PhLP(S) is not. Because PhLP(L) has several consensus sites for the constitutively active kinase casein kinase 2 (CK2) in its N terminus, we tested the phosphorylation of the recombinant proteins by either HEK cell cytosol in the presence or absence of kinase inhibitors or by purified CK2. PhLP(L) was a good CK2 substrate, whereas PhLP(S) and phosducin were not. Progressive truncation and serine/threonine to alanine mutations of the PhLP(L) N terminus identified a serine/threonine cluster (Ser-18/Thr-19/Ser-20) within a small N-terminal region of PhLP(L) (amino acids 5-28) as the site in which PhLP(L) function was modified in HEK 293 cells. In native tissue, PhLP(L) also seems to be regulated by phosphorylation because phosphorylated and non-phosphorylated forms of PhLP(L) were detected in mouse brain and adrenal gland. Moreover, the alternatively spliced isoform PhLP(S) was also found in adrenal tissue. Therefore, the physiological control of G-protein regulation by PhLP seems to involve phosphorylation by CK2 and alternative splicing of the regulator.

@article{Humrich2003, abstract = {Phosducin-like protein (PhLP) is a member of the phosducin family of G-protein betagamma-regulators and exists in two splice variants. The long isoform PhLP(L) and the short isoform PhLP(S) differ by the presence or absence of an 83-amino acid N terminus. In isolated biochemical assay systems, PhLP(L) is the more potent Gbetagamma-inhibitor, whereas the functional role of PhLP(S) is still unclear. We now report that in intact HEK 293 cells, PhLP(S) inhibited Gbetagamma-induced inositol phosphate generation with approximately 20-fold greater potency than PhLP(L). Radiolabeling of transfected HEK 293 cells with [(32)P] revealed that PhLP(L) is constitutively phosphorylated, whereas PhLP(S) is not. Because PhLP(L) has several consensus sites for the constitutively active kinase casein kinase 2 (CK2) in its N terminus, we tested the phosphorylation of the recombinant proteins by either HEK cell cytosol in the presence or absence of kinase inhibitors or by purified CK2. PhLP(L) was a good CK2 substrate, whereas PhLP(S) and phosducin were not. Progressive truncation and serine/threonine to alanine mutations of the PhLP(L) N terminus identified a serine/threonine cluster (Ser-18/Thr-19/Ser-20) within a small N-terminal region of PhLP(L) (amino acids 5-28) as the site in which PhLP(L) function was modified in HEK 293 cells. In native tissue, PhLP(L) also seems to be regulated by phosphorylation because phosphorylated and non-phosphorylated forms of PhLP(L) were detected in mouse brain and adrenal gland. Moreover, the alternatively spliced isoform PhLP(S) was also found in adrenal tissue. Therefore, the physiological control of G-protein regulation by PhLP seems to involve phosphorylation by CK2 and alternative splicing of the regulator.}, added-at = {2010-12-14T18:12:02.000+0100}, author = {Humrich, J. and Bermel, C. and Grubel, T. and Quitterer, U. and Lohse, M. J.}, biburl = {https://www.bibsonomy.org/bibtex/224384e471316b95288949944d8889660/pharmawuerz}, endnotereftype = {Journal Article}, interhash = {547801d073c20878d4fed705d4ae7b58}, intrahash = {24384e471316b95288949944d8889660}, issn = {0021-9258 (Print) 0021-9258 (Linking)}, journal = {J Biol Chem}, keywords = {& Alternative Animals Carrier Casein Cell Enzyme Humans II Inhibitors/pharmacology Kinase Kinases/antagonists Line Mice Nerve Organ Phosphorylation/drug Protein-Serine-Threonine Proteins/genetics/*metabolism Specificity Splicing Substrate Tissue effects inhibitors/genetics/*metabolism}, month = {Feb 14}, note = {Humrich, Jan Bermel, Christina Grubel, Tobias Quitterer, Ursula Lohse, Martin J United States The Journal of biological chemistry J Biol Chem. 2003 Feb 14;278(7):4474-81. Epub 2002 Dec 3.}, number = 7, pages = {4474-81}, shorttitle = {Regulation of phosducin-like protein by casein kinase 2 and N-terminal splicing}, timestamp = {2010-12-14T18:12:15.000+0100}, title = {Regulation of phosducin-like protein by casein kinase 2 and N-terminal splicing}, url = {http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12466282}, volume = 278, year = 2003 }