• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of geneticsGeneticsCurrent IssueInformation for AuthorsEditorial BoardSubscribeSubmit a Manuscript
Genetics. Mar 1999; 151(3): 1065–1079.
PMCID: PMC1460532

Genetic analysis of viable Hsp90 alleles reveals a critical role in Drosophila spermatogenesis.

Abstract

The Hsp90 chaperone protein maintains the activities of a remarkable variety of signal transducers, but its most critical functions in the context of the whole organism are unknown. Point mutations of Hsp83 (the Drosophila Hsp90 gene) obtained in two different screens are lethal as homozygotes. We report that eight transheterozygous mutant combinations produce viable adults. All exhibit the same developmental defects: sterile males and sterile or weakly fertile females. We also report that scratch, a previously identified male-sterile mutation, is an allele of Hsp82 with a P-element insertion in the intron that reduces expression. Thus, it is a simple reduction in Hsp90 function, rather than possible altered functions in the point mutants, that leads to male sterility. As shown by light and electron microscopy, all stages of spermatogenesis involving microtubule function are affected, from early mitotic divisions to later stages of sperm maturation, individualization, and motility. Aberrant microtubules are prominent in yeast cells carrying mutations in HSP82 (the yeast Hsp90 gene), confirming that Hsp90 function is connected to microtubule dynamics and that this connection is highly conserved. A small fraction of Hsp90 copurifies with taxol-stabilized microtubule proteins in Drosophila embryo extracts, but Hsp90 does not remain associated with microtubules through repeated temperature-induced assembly and disassembly reactions. If the spermatogenesis phenotypes are due to defects in microtubule dynamics, we suggest these are indirect, reflecting a role for Hsp90 in maintaining critical signal transduction pathways and microtubule effectors, rather than a direct role in the assembly and disassembly of microtubules themselves.

Full Text

The Full Text of this article is available as a PDF (586K).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Aligue R, Akhavan-Niak H, Russell P. A role for Hsp90 in cell cycle control: Wee1 tyrosine kinase activity requires interaction with Hsp90. EMBO J. 1994 Dec 15;13(24):6099–6106. [PMC free article] [PubMed]
  • Antonsson C, Whitelaw ML, McGuire J, Gustafsson JA, Poellinger L. Distinct roles of the molecular chaperone hsp90 in modulating dioxin receptor function via the basic helix-loop-helix and PAS domains. Mol Cell Biol. 1995 Feb;15(2):756–765. [PMC free article] [PubMed]
  • Nathan Peter A, Meadows Kenneth D, Keniston Richard C, Lockwood Richard S. Predictive value of nerve conduction studies. Occup Environ Med. 1997 Oct;54(10):a765–765. [PMC free article]
  • Baker BS. Paternal loss (pal): a meiotic mutant in Drosophila melanogaster causing loss of paternal chromosomes. Genetics. 1975 Jun;80(2):267–296. [PMC free article] [PubMed]
  • Bardwell JC, Craig EA. Ancient heat shock gene is dispensable. J Bacteriol. 1988 Jul;170(7):2977–2983. [PMC free article] [PubMed]
  • Biggiogera M, Tanguay RM, Marin R, Wu Y, Martin TE, Fakan S. Localization of heat shock proteins in mouse male germ cells: an immunoelectron microscopical study. Exp Cell Res. 1996 Nov 25;229(1):77–85. [PubMed]
  • Borkovich KA, Farrelly FW, Finkelstein DB, Taulien J, Lindquist S. hsp82 is an essential protein that is required in higher concentrations for growth of cells at higher temperatures. Mol Cell Biol. 1989 Sep;9(9):3919–3930. [PMC free article] [PubMed]
  • Bozzetti MP, Massari S, Finelli P, Meggio F, Pinna LA, Boldyreff B, Issinger OG, Palumbo G, Ciriaco C, Bonaccorsi S, et al. The Ste locus, a component of the parasitic cry-Ste system of Drosophila melanogaster, encodes a protein that forms crystals in primary spermatocytes and mimics properties of the beta subunit of casein kinase 2. Proc Natl Acad Sci U S A. 1995 Jun 20;92(13):6067–6071. [PMC free article] [PubMed]
  • Bose S, Weikl T, Bügl H, Buchner J. Chaperone function of Hsp90-associated proteins. Science. 1996 Dec 6;274(5293):1715–1717. [PubMed]
  • Bozzetti MP, Massari S, Finelli P, Meggio F, Pinna LA, Boldyreff B, Issinger OG, Palumbo G, Ciriaco C, Bonaccorsi S, et al. The Ste locus, a component of the parasitic cry-Ste system of Drosophila melanogaster, encodes a protein that forms crystals in primary spermatocytes and mimics properties of the beta subunit of casein kinase 2. Proc Natl Acad Sci U S A. 1995 Jun 20;92(13):6067–6071. [PMC free article] [PubMed]
  • Brugge JS. Interaction of the Rous sarcoma virus protein pp60src with the cellular proteins pp50 and pp90. Curr Top Microbiol Immunol. 1986;123:1–22. [PubMed]
  • Carbajal ME, Valet JP, Charest PM, Tanguay RM. Purification of Drosophila hsp 83 and immunoelectron microscopic localization. Eur J Cell Biol. 1990 Jun;52(1):147–156. [PubMed]
  • Castrillon DH, Gönczy P, Alexander S, Rawson R, Eberhart CG, Viswanathan S, DiNardo S, Wasserman SA. Toward a molecular genetic analysis of spermatogenesis in Drosophila melanogaster: characterization of male-sterile mutants generated by single P element mutagenesis. Genetics. 1993 Oct;135(2):489–505. [PMC free article] [PubMed]
  • Cooley L, Kelley R, Spradling A. Insertional mutagenesis of the Drosophila genome with single P elements. Science. 1988 Mar 4;239(4844):1121–1128. [PubMed]
  • Cutforth T, Rubin GM. Mutations in Hsp83 and cdc37 impair signaling by the sevenless receptor tyrosine kinase in Drosophila. Cell. 1994 Jul 1;77(7):1027–1036. [PubMed]
  • Czar MJ, Welsh MJ, Pratt WB. Immunofluorescence localization of the 90-kDa heat-shock protein to cytoskeleton. Eur J Cell Biol. 1996 Aug;70(4):322–330. [PubMed]
  • Dey B, Caplan AJ, Boschelli F. The Ydj1 molecular chaperone facilitates formation of active p60v-src in yeast. Mol Biol Cell. 1996 Jan;7(1):91–100. [PMC free article] [PubMed]
  • Dickson BJ, van der Straten A, Dominguez M, Hafen E. Mutations Modulating Raf signaling in Drosophila eye development. Genetics. 1996 Jan;142(1):163–171. [PMC free article] [PubMed]
  • Enan E, Matsumura F. Evidence for a second pathway in the action mechanism of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Significance of Ah-receptor mediated activation of protein kinase under cell-free conditions. Biochem Pharmacol. 1995 Jan 18;49(2):249–261. [PubMed]
  • Fostinis Y, Theodoropoulos PA, Gravanis A, Stournaras C. Heat shock protein HSP90 and its association with the cytoskeleton: a morphological study. Biochem Cell Biol. 1992 Sep;70(9):779–786. [PubMed]
  • Minami Y, Kawasaki H, Suzuki K, Yahara I. The calmodulin-binding domain of the mouse 90-kDa heat shock protein. J Biol Chem. 1993 May 5;268(13):9604–9610. [PubMed]
  • Minami Y, Kimura Y, Kawasaki H, Suzuki K, Yahara I. The carboxy-terminal region of mammalian HSP90 is required for its dimerization and function in vivo. Mol Cell Biol. 1994 Feb;14(2):1459–1464. [PMC free article] [PubMed]
  • Miyata Y, Yahara I. The 90-kDa heat shock protein, HSP90, binds and protects casein kinase II from self-aggregation and enhances its kinase activity. J Biol Chem. 1992 Apr 5;267(10):7042–7047. [PubMed]
  • Fuyama Y. Genetics of Parthenogenesis in DROSOPHILA MELANOGASTER. I. the Modes of Diploidization in the Gynogenesis Induced by a Male-Sterile Mutant, ms(3)K81. Genetics. 1986 Feb;112(2):237–248. [PMC free article] [PubMed]
  • Miyata Y, Yahara I. Interaction between casein kinase II and the 90-kDa stress protein, HSP90. Biochemistry. 1995 Jun 27;34(25):8123–8129. [PubMed]
  • Goldstein LS, Laymon RA, McIntosh JR. A microtubule-associated protein in Drosophila melanogaster: identification, characterization, and isolation of coding sequences. J Cell Biol. 1986 Jun;102(6):2076–2087. [PMC free article] [PubMed]
  • Nathan DF, Lindquist S. Mutational analysis of Hsp90 function: interactions with a steroid receptor and a protein kinase. Mol Cell Biol. 1995 Jul;15(7):3917–3925. [PMC free article] [PubMed]
  • Graner M, Stupka K, Karr TL. Biochemical and cytological characterization of DROP-1: a widely distributed proteoglycan in Drosophila. Insect Biochem Mol Biol. 1994 Jun;24(6):557–567. [PubMed]
  • Nishida E, Koyasu S, Sakai H, Yahara I. Calmodulin-regulated binding of the 90-kDa heat shock protein to actin filaments. J Biol Chem. 1986 Dec 5;261(34):16033–16036. [PubMed]
  • Holley SJ, Yamamoto KR. A role for Hsp90 in retinoid receptor signal transduction. Mol Biol Cell. 1995 Dec;6(12):1833–1842. [PMC free article] [PubMed]
  • Nordeen SK, Moyer ML, Bona BJ. The coupling of multiple signal transduction pathways with steroid response mechanisms. Endocrinology. 1994 Apr;134(4):1723–1732. [PubMed]
  • Holmgren R, Livak K, Morimoto R, Freund R, Meselson M. Studies of cloned sequences from four Drosophila heat shock loci. Cell. 1979 Dec;18(4):1359–1370. [PubMed]
  • Ohsako S, Bunick D, Hayashi Y. Immunocytochemical observation of the 90 KD heat shock protein (HSP90): high expression in primordial and pre-meiotic germ cells of male and female rat gonads. J Histochem Cytochem. 1995 Jan;43(1):67–76. [PubMed]
  • Jaiswal RK, Weissinger E, Kolch W, Landreth GE. Nerve growth factor-mediated activation of the mitogen-activated protein (MAP) kinase cascade involves a signaling complex containing B-Raf and HSP90. J Biol Chem. 1996 Sep 27;271(39):23626–23629. [PubMed]
  • Olmsted JB, Borisy GG. Characterization of microtubule assembly in porcine brain extracts by viscometry. Biochemistry. 1973 Oct 9;12(21):4282–4289. [PubMed]
  • Jakob U, Lilie H, Meyer I, Buchner J. Transient interaction of Hsp90 with early unfolding intermediates of citrate synthase. Implications for heat shock in vivo. J Biol Chem. 1995 Mar 31;270(13):7288–7294. [PubMed]
  • Palumbo G, Berloco M, Fanti L, Bozzetti MP, Massari S, Caizzi R, Caggese C, Spinelli L, Pimpinelli S. Interaction systems between heterochromatin and euchromatin in Drosophila melanogaster. Genetica. 1994;94(2-3):267–274. [PubMed]
  • Jaskoll T, Choy HA, Melnick M. The glucocorticoid-glucocorticoid receptor signal transduction pathway, transforming growth factor-beta, and embryonic mouse lung development in vivo. Pediatr Res. 1996 May;39(5):749–759. [PubMed]
  • Karpen GH, Spradling AC. Analysis of subtelomeric heterochromatin in the Drosophila minichromosome Dp1187 by single P element insertional mutagenesis. Genetics. 1992 Nov;132(3):737–753. [PMC free article] [PubMed]
  • Perdew GH, Hord N, Hollenback CE, Welsh MJ. Localization and characterization of the 86- and 84-kDa heat shock proteins in Hepa 1c1c7 cells. Exp Cell Res. 1993 Dec;209(2):350–356. [PubMed]
  • Karr TL. Paternal investment and intracellular sperm-egg interactions during and following fertilization in Drosophila. Curr Top Dev Biol. 1996;34:89–115. [PubMed]
  • Perrot-Applanat M, Cibert C, Géraud G, Renoir JM, Baulieu EE. The 59 kDa FK506-binding protein, a 90 kDa heat shock protein binding immunophilin (FKBP59-HBI), is associated with the nucleus, the cytoskeleton and mitotic apparatus. J Cell Sci. 1995 May;108(Pt 5):2037–2051. [PubMed]
  • Kemphues KJ, Raff EC, Kaufman TC. Genetic analysis of B2t, the structural gene for a testis-specific beta-tubulin subunit in Drosophila melanogaster. Genetics. 1983 Oct;105(2):345–356. [PMC free article] [PubMed]
  • Kimura Y, Yahara I, Lindquist S. Role of the protein chaperone YDJ1 in establishing Hsp90-mediated signal transduction pathways. Science. 1995 Jun 2;268(5215):1362–1365. [PubMed]
  • Pettersson K, Svensson K, Mattsson R, Carlsson B, Ohlsson R, Berkenstam A. Expression of a novel member of estrogen response element-binding nuclear receptors is restricted to the early stages of chorion formation during mouse embryogenesis. Mech Dev. 1996 Feb;54(2):211–223. [PubMed]
  • Kimura Y, Rutherford SL, Miyata Y, Yahara I, Freeman BC, Yue L, Morimoto RI, Lindquist S. Cdc37 is a molecular chaperone with specific functions in signal transduction. Genes Dev. 1997 Jul 15;11(14):1775–1785. [PubMed]
  • Picard D, Khursheed B, Garabedian MJ, Fortin MG, Lindquist S, Yamamoto KR. Reduced levels of hsp90 compromise steroid receptor action in vivo. Nature. 1990 Nov 8;348(6297):166–168. [PubMed]
  • Lifschytz E. Uncoupling of gonial and spermatocyte stages by means of conditional lethal mutations in Drosophila melanogaster. Dev Biol. 1978 Oct;66(2):571–578. [PubMed]
  • Pimpinelli S, Sullivan W, Prout M, Sandler L. On biological functions mapping to the heterochromatin of Drosophila melanogaster. Genetics. 1985 Apr;109(4):701–724. [PMC free article] [PubMed]
  • Lifschytz E. The developmental program of spermiogenesis in Drosophila: a genetic analysis. Int Rev Cytol. 1987;109:211–258. [PubMed]
  • Pitnick S, Spicer GS, Markow TA. How long is a giant sperm? Nature. 1995 May 11;375(6527):109–109. [PubMed]
  • Pongratz I, Mason GG, Poellinger L. Dual roles of the 90-kDa heat shock protein hsp90 in modulating functional activities of the dioxin receptor. Evidence that the dioxin receptor functionally belongs to a subclass of nuclear receptors which require hsp90 both for ligand binding activity and repression of intrinsic DNA binding activity. J Biol Chem. 1992 Jul 5;267(19):13728–13734. [PubMed]
  • Livak KJ. Organization and mapping of a sequence on the Drosophila melanogaster X and Y chromosomes that is transcribed during spermatogenesis. Genetics. 1984 Aug;107(4):611–634. [PMC free article] [PubMed]
  • Pratt WB. Control of steroid receptor function and cytoplasmic-nuclear transport by heat shock proteins. Bioessays. 1992 Dec;14(12):841–848. [PubMed]
  • Louvion JF, Warth R, Picard D. Two eukaryote-specific regions of Hsp82 are dispensable for its viability and signal transduction functions in yeast. Proc Natl Acad Sci U S A. 1996 Nov 26;93(24):13937–13942. [PMC free article] [PubMed]
  • Pratt WB, Toft DO. Steroid receptor interactions with heat shock protein and immunophilin chaperones. Endocr Rev. 1997 Jun;18(3):306–360. [PubMed]
  • Pringle JR, Adams AE, Drubin DG, Haarer BK. Immunofluorescence methods for yeast. Methods Enzymol. 1991;194:565–602. [PubMed]
  • Stebbins CE, Russo AA, Schneider C, Rosen N, Hartl FU, Pavletich NP. Crystal structure of an Hsp90-geldanamycin complex: targeting of a protein chaperone by an antitumor agent. Cell. 1997 Apr 18;89(2):239–250. [PubMed]
  • Prodromou C, Roe SM, O'Brien R, Ladbury JE, Piper PW, Pearl LH. Identification and structural characterization of the ATP/ADP-binding site in the Hsp90 molecular chaperone. Cell. 1997 Jul 11;90(1):65–75. [PubMed]
  • van der Straten A, Rommel C, Dickson B, Hafen E. The heat shock protein 83 (Hsp83) is required for Raf-mediated signalling in Drosophila. EMBO J. 1997 Apr 15;16(8):1961–1969. [PMC free article] [PubMed]
  • Prodromou C, Roe SM, Piper PW, Pearl LH. A molecular clamp in the crystal structure of the N-terminal domain of the yeast Hsp90 chaperone. Nat Struct Biol. 1997 Jun;4(6):477–482. [PubMed]
  • Velazquez JM, Sonoda S, Bugaisky G, Lindquist S. Is the major Drosophila heat shock protein present in cells that have not been heat shocked? J Cell Biol. 1983 Jan;96(1):286–290. [PMC free article] [PubMed]
  • Redmond T, Sanchez ER, Bresnick EH, Schlesinger MJ, Toft DO, Pratt WB, Welsh MJ. Immunofluorescence colocalization of the 90-kDa heat-shock protein and microtubules in interphase and mitotic mammalian cells. Eur J Cell Biol. 1989 Oct;50(1):66–75. [PubMed]
  • Wearsch PA, Nicchitta CV. Endoplasmic reticulum chaperone GRP94 subunit assembly is regulated through a defined oligomerization domain. Biochemistry. 1996 Dec 24;35(51):16760–16769. [PubMed]
  • Rose DW, Welch WJ, Kramer G, Hardesty B. Possible involvement of the 90-kDa heat shock protein in the regulation of protein synthesis. J Biol Chem. 1989 Apr 15;264(11):6239–6244. [PubMed]
  • Rutherford SL, Lindquist S. Hsp90 as a capacitor for morphological evolution. Nature. 1998 Nov 26;396(6709):336–342. [PubMed]
  • Sanchez ER, Redmond T, Scherrer LC, Bresnick EH, Welsh MJ, Pratt WB. Evidence that the 90-kilodalton heat shock protein is associated with tubulin-containing complexes in L cell cytosol and in intact PtK cells. Mol Endocrinol. 1988 Aug;2(8):756–760. [PubMed]
  • Xu Y, Lindquist S. Heat-shock protein hsp90 governs the activity of pp60v-src kinase. Proc Natl Acad Sci U S A. 1993 Aug 1;90(15):7074–7078. [PMC free article] [PubMed]
  • Xu Y, Singer MA, Lindquist S. Maturation of the tyrosine kinase c-src as a kinase and as a substrate depends on the molecular chaperone Hsp90. Proc Natl Acad Sci U S A. 1999 Jan 5;96(1):109–114. [PMC free article] [PubMed]
  • Yasuda GK, Schubiger G, Wakimoto BT. Genetic characterization of ms (3) K81, a paternal effect gene of Drosophila melanogaster. Genetics. 1995 May;140(1):219–229. [PMC free article] [PubMed]
  • Srinivasan S, Karr TL. Biochemical characterization of related microtubule proteins in Drosophila melanogaster and adult rat brain. Brain Res. 1995 Dec 1;701(1-2):39–46. [PubMed]
  • Yue L, Spradling AC. hu-li tai shao, a gene required for ring canal formation during Drosophila oogenesis, encodes a homolog of adducin. Genes Dev. 1992 Dec;6(12B):2443–2454. [PubMed]
  • Stancato LF, Chow YH, Hutchison KA, Perdew GH, Jove R, Pratt WB. Raf exists in a native heterocomplex with hsp90 and p50 that can be reconstituted in a cell-free system. J Biol Chem. 1993 Oct 15;268(29):21711–21716. [PubMed]

Articles from Genetics are provided here courtesy of Genetics Society of America

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...