Abstract
The alpha-crystallins (alpha A and alpha B) are major water-soluble proteins of the transparent eye lens that are expressed in a variety of tissues and can function as molecular chaperones. alpha B-crystallin is also a small heat shock protein associated with numerous degenerative diseases and abnormal growth patterns. Previous experiments have shown that alpha A-and alpha B-crystallin are phosphorylated on specific serine residues by a cAMP-dependent pathway. Here we provide evidence that either total bovine alpha-crystallin or its isolated polypeptides can autophosphorylate serine by a cAMP-independent mechanism in the presence of Mg2+ and [gamma-32P]ATP; the autophosphorylated products isoelectrically focus with the authentic phosphorylated forms of the alpha-crystallin polypeptides. Thus, the alpha A- and alpha B-crystallin/small heat shock protein polypeptides are enzyme-crystallins which may be involved in metabolic pathways important for the development, maintenance, or pathology of the lens and other tissues.
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- Aoyama A., Fröhli E., Schäfer R., Klemenz R. Alpha B-crystallin expression in mouse NIH 3T3 fibroblasts: glucocorticoid responsiveness and involvement in thermal protection. Mol Cell Biol. 1993 Mar;13(3):1824–1835. doi: 10.1128/mcb.13.3.1824. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Atomi Y., Yamada S., Strohman R., Nonomura Y. Alpha B-crystallin in skeletal muscle: purification and localization. J Biochem. 1991 Nov;110(5):812–822. doi: 10.1093/oxfordjournals.jbchem.a123665. [DOI] [PubMed] [Google Scholar]
- Bhat S. P., Nagineni C. N. alpha B subunit of lens-specific protein alpha-crystallin is present in other ocular and non-ocular tissues. Biochem Biophys Res Commun. 1989 Jan 16;158(1):319–325. doi: 10.1016/s0006-291x(89)80215-3. [DOI] [PubMed] [Google Scholar]
- Bloemendal H., de Jong W. W. Lens proteins and their genes. Prog Nucleic Acid Res Mol Biol. 1991;41:259–281. doi: 10.1016/s0079-6603(08)60012-4. [DOI] [PubMed] [Google Scholar]
- Chiesa R., Gawinowicz-Kolks M. A., Spector A. The phosphorylation of the primary gene products of alpha-crystallin. J Biol Chem. 1987 Feb 5;262(4):1438–1441. [PubMed] [Google Scholar]
- Chiesa R., Spector A. The dephosphorylation of lens alpha-crystallin A chain. Biochem Biophys Res Commun. 1989 Aug 15;162(3):1494–1501. doi: 10.1016/0006-291x(89)90843-7. [DOI] [PubMed] [Google Scholar]
- Craig E. A., Gross C. A. Is hsp70 the cellular thermometer? Trends Biochem Sci. 1991 Apr;16(4):135–140. doi: 10.1016/0968-0004(91)90055-z. [DOI] [PubMed] [Google Scholar]
- Crête P., Landry J. Induction of HSP27 phosphorylation and thermoresistance in Chinese hamster cells by arsenite, cycloheximide, A23187, and EGTA. Radiat Res. 1990 Mar;121(3):320–327. [PubMed] [Google Scholar]
- Csermely P., Kahn C. R. The 90-kDa heat shock protein (hsp-90) possesses an ATP binding site and autophosphorylating activity. J Biol Chem. 1991 Mar 15;266(8):4943–4950. [PubMed] [Google Scholar]
- Dasgupta S., Hohman T. C., Carper D. Hypertonic stress induces alpha B-crystallin expression. Exp Eye Res. 1992 Mar;54(3):461–470. doi: 10.1016/0014-4835(92)90058-z. [DOI] [PubMed] [Google Scholar]
- Dubin R. A., Wawrousek E. F., Piatigorsky J. Expression of the murine alpha B-crystallin gene is not restricted to the lens. Mol Cell Biol. 1989 Mar;9(3):1083–1091. doi: 10.1128/mcb.9.3.1083. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Duguid J. R., Rohwer R. G., Seed B. Isolation of cDNAs of scrapie-modulated RNAs by subtractive hybridization of a cDNA library. Proc Natl Acad Sci U S A. 1988 Aug;85(15):5738–5742. doi: 10.1073/pnas.85.15.5738. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Freiden P. J., Gaut J. R., Hendershot L. M. Interconversion of three differentially modified and assembled forms of BiP. EMBO J. 1992 Jan;11(1):63–70. doi: 10.1002/j.1460-2075.1992.tb05028.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gaestel M., Schröder W., Benndorf R., Lippmann C., Buchner K., Hucho F., Erdmann V. A., Bielka H. Identification of the phosphorylation sites of the murine small heat shock protein hsp25. J Biol Chem. 1991 Aug 5;266(22):14721–14724. [PubMed] [Google Scholar]
- Gamer J., Bujard H., Bukau B. Physical interaction between heat shock proteins DnaK, DnaJ, and GrpE and the bacterial heat shock transcription factor sigma 32. Cell. 1992 May 29;69(5):833–842. doi: 10.1016/0092-8674(92)90294-m. [DOI] [PubMed] [Google Scholar]
- Gaut J. R., Hendershot L. M. The immunoglobulin-binding protein in vitro autophosphorylation site maps to a threonine within the ATP binding cleft but is not a detectable site of in vivo phosphorylation. J Biol Chem. 1993 Jun 15;268(17):12691–12698. [PubMed] [Google Scholar]
- Gutkind J. S., Robbins K. C. Translocation of the FGR protein-tyrosine kinase as a consequence of neutrophil activation. Proc Natl Acad Sci U S A. 1989 Nov;86(22):8783–8787. doi: 10.1073/pnas.86.22.8783. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hanks S. K., Quinn A. M. Protein kinase catalytic domain sequence database: identification of conserved features of primary structure and classification of family members. Methods Enzymol. 1991;200:38–62. doi: 10.1016/0076-6879(91)00126-h. [DOI] [PubMed] [Google Scholar]
- Horwitz J. Alpha-crystallin can function as a molecular chaperone. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10449–10453. doi: 10.1073/pnas.89.21.10449. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ingolia T. D., Craig E. A. Four small Drosophila heat shock proteins are related to each other and to mammalian alpha-crystallin. Proc Natl Acad Sci U S A. 1982 Apr;79(7):2360–2364. doi: 10.1073/pnas.79.7.2360. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Iwaki T., Iwaki A., Liem R. K., Goldman J. E. Expression of alpha B-crystallin in the developing rat kidney. Kidney Int. 1991 Jul;40(1):52–56. doi: 10.1038/ki.1991.178. [DOI] [PubMed] [Google Scholar]
- Iwaki T., Iwaki A., Miyazono M., Goldman J. E. Preferential expression of alpha B-crystallin in astrocytic elements of neuroectodermal tumors. Cancer. 1991 Nov 15;68(10):2230–2240. doi: 10.1002/1097-0142(19911115)68:10<2230::aid-cncr2820681023>3.0.co;2-7. [DOI] [PubMed] [Google Scholar]
- Iwaki T., Kume-Iwaki A., Liem R. K., Goldman J. E. Alpha B-crystallin is expressed in non-lenticular tissues and accumulates in Alexander's disease brain. Cell. 1989 Apr 7;57(1):71–78. doi: 10.1016/0092-8674(89)90173-6. [DOI] [PubMed] [Google Scholar]
- Iwaki T., Tateishi J. Immunohistochemical demonstration of alphaB-crystallin in hamartomas of tuberous sclerosis. Am J Pathol. 1991 Dec;139(6):1303–1308. [PMC free article] [PubMed] [Google Scholar]
- Iwaki T., Wisniewski T., Iwaki A., Corbin E., Tomokane N., Tateishi J., Goldman J. E. Accumulation of alpha B-crystallin in central nervous system glia and neurons in pathologic conditions. Am J Pathol. 1992 Feb;140(2):345–356. [PMC free article] [PubMed] [Google Scholar]
- Jakob U., Gaestel M., Engel K., Buchner J. Small heat shock proteins are molecular chaperones. J Biol Chem. 1993 Jan 25;268(3):1517–1520. [PubMed] [Google Scholar]
- Kato K., Shinohara H., Goto S., Inaguma Y., Morishita R., Asano T. Copurification of small heat shock protein with alpha B crystallin from human skeletal muscle. J Biol Chem. 1992 Apr 15;267(11):7718–7725. [PubMed] [Google Scholar]
- Kato K., Shinohara H., Kurobe N., Inaguma Y., Shimizu K., Ohshima K. Tissue distribution and developmental profiles of immunoreactive alpha B crystallin in the rat determined with a sensitive immunoassay system. Biochim Biophys Acta. 1991 May 24;1074(1):201–208. doi: 10.1016/0304-4165(91)90062-l. [DOI] [PubMed] [Google Scholar]
- Kato S., Hirano A., Umahara T., Llena J. F., Herz F., Ohama E. Ultrastructural and immunohistochemical studies on ballooned cortical neurons in Creutzfeldt-Jakob disease: expression of alpha B-crystallin, ubiquitin and stress-response protein 27. Acta Neuropathol. 1992;84(4):443–448. doi: 10.1007/BF00227673. [DOI] [PubMed] [Google Scholar]
- Klemenz R., Andres A. C., Fröhli E., Schäfer R., Aoyama A. Expression of the murine small heat shock proteins hsp 25 and alpha B crystallin in the absence of stress. J Cell Biol. 1993 Feb;120(3):639–645. doi: 10.1083/jcb.120.3.639. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Klemenz R., Fröhli E., Aoyama A., Hoffmann S., Simpson R. J., Moritz R. L., Schäfer R. Alpha B crystallin accumulation is a specific response to Ha-ras and v-mos oncogene expression in mouse NIH 3T3 fibroblasts. Mol Cell Biol. 1991 Feb;11(2):803–812. doi: 10.1128/mcb.11.2.803. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Klemenz R., Fröhli E., Steiger R. H., Schäfer R., Aoyama A. Alpha B-crystallin is a small heat shock protein. Proc Natl Acad Sci U S A. 1991 May 1;88(9):3652–3656. doi: 10.1073/pnas.88.9.3652. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Landry J., Chrétien P., Laszlo A., Lambert H. Phosphorylation of HSP27 during development and decay of thermotolerance in Chinese hamster cells. J Cell Physiol. 1991 Apr;147(1):93–101. doi: 10.1002/jcp.1041470113. [DOI] [PubMed] [Google Scholar]
- Landry J., Lambert H., Zhou M., Lavoie J. N., Hickey E., Weber L. A., Anderson C. W. Human HSP27 is phosphorylated at serines 78 and 82 by heat shock and mitogen-activated kinases that recognize the same amino acid motif as S6 kinase II. J Biol Chem. 1992 Jan 15;267(2):794–803. [PubMed] [Google Scholar]
- Lee D. C., Kim R. Y., Wistow G. J. An avian alpha B-crystallin. Non-lens expression and sequence similarities with both small (HSP27) and large (HSP70) heat shock proteins. J Mol Biol. 1993 Aug 20;232(4):1221–1226. doi: 10.1006/jmbi.1993.1476. [DOI] [PubMed] [Google Scholar]
- Longoni S., James P., Chiesi M. Cardiac alpha-crystallin. I. Isolation and identification. Mol Cell Biochem. 1990 Dec 3;99(1):113–120. [PubMed] [Google Scholar]
- Lowe J., Landon M., Pike I., Spendlove I., McDermott H., Mayer R. J. Dementia with beta-amyloid deposition: involvement of alpha B-crystallin supports two main diseases. Lancet. 1990 Aug 25;336(8713):515–516. doi: 10.1016/0140-6736(90)92075-s. [DOI] [PubMed] [Google Scholar]
- Lowe J., McDermott H., Pike I., Spendlove I., Landon M., Mayer R. J. alpha B crystallin expression in non-lenticular tissues and selective presence in ubiquitinated inclusion bodies in human disease. J Pathol. 1992 Jan;166(1):61–68. doi: 10.1002/path.1711660110. [DOI] [PubMed] [Google Scholar]
- Maiti M., Kono M., Chakrabarti B. Heat-induced changes in the conformation of alpha- and beta-crystallins: unique thermal stability of alpha-crystallin. FEBS Lett. 1988 Aug 15;236(1):109–114. doi: 10.1016/0014-5793(88)80295-3. [DOI] [PubMed] [Google Scholar]
- McCarty J. S., Walker G. C. DnaK as a thermometer: threonine-199 is site of autophosphorylation and is critical for ATPase activity. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9513–9517. doi: 10.1073/pnas.88.21.9513. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Mulders J. W., Stokkermans J., Leunissen J. A., Benedetti E. L., Bloemendal H., de Jong W. W. Interaction of alpha-crystallin with lens plasma membranes. Affinity for MP26. Eur J Biochem. 1985 Nov 4;152(3):721–728. doi: 10.1111/j.1432-1033.1985.tb09253.x. [DOI] [PubMed] [Google Scholar]
- Murano S., Thweatt R., Shmookler Reis R. J., Jones R. A., Moerman E. J., Goldstein S. Diverse gene sequences are overexpressed in werner syndrome fibroblasts undergoing premature replicative senescence. Mol Cell Biol. 1991 Aug;11(8):3905–3914. doi: 10.1128/mcb.11.8.3905. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nadeau K., Das A., Walsh C. T. Hsp90 chaperonins possess ATPase activity and bind heat shock transcription factors and peptidyl prolyl isomerases. J Biol Chem. 1993 Jan 15;268(2):1479–1487. [PubMed] [Google Scholar]
- Piatigorsky J. Lens crystallins. Innovation associated with changes in gene regulation. J Biol Chem. 1992 Mar 5;267(7):4277–4280. [PubMed] [Google Scholar]
- Piatigorsky J. The twelfth Frederick H. Verhoeff Lecture: gene sharing in the visual system. Trans Am Ophthalmol Soc. 1993;91:283–297. [PMC free article] [PubMed] [Google Scholar]
- Piatigorsky J., Wistow G. J. Enzyme/crystallins: gene sharing as an evolutionary strategy. Cell. 1989 Apr 21;57(2):197–199. doi: 10.1016/0092-8674(89)90956-2. [DOI] [PubMed] [Google Scholar]
- Picard D., Khursheed B., Garabedian M. J., Fortin M. G., Lindquist S., Yamamoto K. R. Reduced levels of hsp90 compromise steroid receptor action in vivo. Nature. 1990 Nov 8;348(6297):166–168. doi: 10.1038/348166a0. [DOI] [PubMed] [Google Scholar]
- Renkawek K., de Jong W. W., Merck K. B., Frenken C. W., van Workum F. P., Bosman G. J. alpha B-crystallin is present in reactive glia in Creutzfeldt-Jakob disease. Acta Neuropathol. 1992;83(3):324–327. doi: 10.1007/BF00296796. [DOI] [PubMed] [Google Scholar]
- Roquemore E. P., Dell A., Morris H. R., Panico M., Reason A. J., Savoy L. A., Wistow G. J., Zigler J. S., Jr, Earles B. J., Hart G. W. Vertebrate lens alpha-crystallins are modified by O-linked N-acetylglucosamine. J Biol Chem. 1992 Jan 5;267(1):555–563. [PubMed] [Google Scholar]
- Russell P., Yamada T. Increased sensitivity of two-dimensional gel electrophoresis using PhastSystem. Biotechniques. 1990 Oct;9(4):422–422,424. [PubMed] [Google Scholar]
- Schieven G., Martin G. S. Nonenzymatic phosphorylation of tyrosine and serine by ATP is catalyzed by manganese but not magnesium. J Biol Chem. 1988 Oct 25;263(30):15590–15593. [PubMed] [Google Scholar]
- Scotting P., McDermott H., Mayer R. J. Ubiquitin-protein conjugates and alpha B crystallin are selectively present in cells undergoing major cytomorphological reorganisation in early chicken embryos. FEBS Lett. 1991 Jul 8;285(1):75–79. doi: 10.1016/0014-5793(91)80728-l. [DOI] [PubMed] [Google Scholar]
- Sherman MYu, Goldberg A. L. Heat shock in Escherichia coli alters the protein-binding properties of the chaperonin groEL by inducing its phosphorylation. Nature. 1992 May 14;357(6374):167–169. doi: 10.1038/357167a0. [DOI] [PubMed] [Google Scholar]
- Sherman MYu, Goldberg A. L. Involvement of the chaperonin dnaK in the rapid degradation of a mutant protein in Escherichia coli. EMBO J. 1992 Jan;11(1):71–77. doi: 10.1002/j.1460-2075.1992.tb05029.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sherman M. Y., Goldberg A. L. Heat shock of Escherichia coli increases binding of dnaK (the hsp70 homolog) to polypeptides by promoting its phosphorylation. Proc Natl Acad Sci U S A. 1993 Sep 15;90(18):8648–8652. doi: 10.1073/pnas.90.18.8648. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Spector A., Chiesa R., Sredy J., Garner W. cAMP-dependent phosphorylation of bovine lens alpha-crystallin. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4712–4716. doi: 10.1073/pnas.82.14.4712. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sredy J., Spector A. The phosphorylation of bovine and human lens polypeptides. Exp Eye Res. 1984 Nov;39(5):653–664. doi: 10.1016/0014-4835(84)90064-2. [DOI] [PubMed] [Google Scholar]
- Srinivasan A. N., Nagineni C. N., Bhat S. P. alpha A-crystallin is expressed in non-ocular tissues. J Biol Chem. 1992 Nov 15;267(32):23337–23341. [PubMed] [Google Scholar]
- Straus D. B., Walter W. A., Gross C. A. Escherichia coli heat shock gene mutants are defective in proteolysis. Genes Dev. 1988 Dec;2(12B):1851–1858. doi: 10.1101/gad.2.12b.1851. [DOI] [PubMed] [Google Scholar]
- Tomokane N., Iwaki T., Tateishi J., Iwaki A., Goldman J. E. Rosenthal fibers share epitopes with alpha B-crystallin, glial fibrillary acidic protein, and ubiquitin, but not with vimentin. Immunoelectron microscopy with colloidal gold. Am J Pathol. 1991 Apr;138(4):875–885. [PMC free article] [PubMed] [Google Scholar]
- Voorter C. E., Mulders J. W., Bloemendal H., de Jong W. W. Some aspects of the phosphorylation of alpha-crystallin A. Eur J Biochem. 1986 Oct 1;160(1):203–210. doi: 10.1111/j.1432-1033.1986.tb09958.x. [DOI] [PubMed] [Google Scholar]
- Whitehouse S., Walsh D. A. Inhibitor protein of the cAMP-dependent protein kinase: characteristics and purification. Methods Enzymol. 1983;99:80–93. doi: 10.1016/0076-6879(83)99044-4. [DOI] [PubMed] [Google Scholar]
- Wickner S. H. Three Escherichia coli heat shock proteins are required for P1 plasmid DNA replication: formation of an active complex between E. coli DnaJ protein and the P1 initiator protein. Proc Natl Acad Sci U S A. 1990 Apr;87(7):2690–2694. doi: 10.1073/pnas.87.7.2690. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wistow G. J., Piatigorsky J. Lens crystallins: the evolution and expression of proteins for a highly specialized tissue. Annu Rev Biochem. 1988;57:479–504. doi: 10.1146/annurev.bi.57.070188.002403. [DOI] [PubMed] [Google Scholar]
- Wistow G. Evolution of a protein superfamily: relationships between vertebrate lens crystallins and microorganism dormancy proteins. J Mol Evol. 1990 Feb;30(2):140–145. doi: 10.1007/BF02099940. [DOI] [PubMed] [Google Scholar]
- Zantema A., Verlaan-De Vries M., Maasdam D., Bol S., van der Eb A. Heat shock protein 27 and alpha B-crystallin can form a complex, which dissociates by heat shock. J Biol Chem. 1992 Jun 25;267(18):12936–12941. [PubMed] [Google Scholar]
- Zhou M., Lambert H., Landry J. Transient activation of a distinct serine protein kinase is responsible for 27-kDa heat shock protein phosphorylation in mitogen-stimulated and heat-shocked cells. J Biol Chem. 1993 Jan 5;268(1):35–43. [PubMed] [Google Scholar]
- Zylicz M., LeBowitz J. H., McMacken R., Georgopoulos C. The dnaK protein of Escherichia coli possesses an ATPase and autophosphorylating activity and is essential in an in vitro DNA replication system. Proc Natl Acad Sci U S A. 1983 Nov;80(21):6431–6435. doi: 10.1073/pnas.80.21.6431. [DOI] [PMC free article] [PubMed] [Google Scholar]
- de Jong W. W., Hendriks W., Mulders J. W., Bloemendal H. Evolution of eye lens crystallins: the stress connection. Trends Biochem Sci. 1989 Sep;14(9):365–368. doi: 10.1016/0968-0004(89)90009-1. [DOI] [PubMed] [Google Scholar]
- de Jong W. W., Leunissen J. A., Voorter C. E. Evolution of the alpha-crystallin/small heat-shock protein family. Mol Biol Evol. 1993 Jan;10(1):103–126. doi: 10.1093/oxfordjournals.molbev.a039992. [DOI] [PubMed] [Google Scholar]