title: The multiple functions of cysteine-string protein analyzed at Drosophila nerve terminals.
abstract: The synaptic vesicle-associated cysteine-string protein (CSP) is important for synaptic transmission. Previous studies revealed multiple defects at neuromuscular junctions (NMJs) of csp null-mutant Drosophila, but whether these defects are independent of each other or mechanistically linked through J domain mediated-interactions with heat-shock cognate protein 70 (Hsc70) has not been established. To resolve this issue, we genetically dissected the individual functions of CSP by an in vivo structure/function analysis. Expression of mutant CSP lacking the J domain at csp null-mutant NMJs fully restored normal thermo-tolerance of evoked transmitter release but did not completely restore evoked release at room temperature and failed to reverse the abnormal intraterminal Ca2+ levels. This suggests that J domain-mediated functions are essential for the regulation of intraterminal Ca2+ levels but only partially required for regulating evoked release and not required for protecting evoked release against thermal stress. Hence, CSP can also act as an Hsc70-independent chaperone protecting evoked release from thermal stress. Expression of mutant CSP lacking the L domain restored neurotransmission and partially reversed the abnormal intraterminal Ca2+ levels, suggesting that the L domain is important, although not essential, for the role of CSP in regulating intraterminal Ca2+ levels. We detected no effects of csp mutations on individual presynaptic Ca2+ signals triggered by action potentials, suggesting that presynaptic Ca2+ entry is not primarily impaired. Both the J and L domains were also required for the role of CSP in synaptic growth. Together, these results suggest that CSP has several independent synaptic functions, affecting synaptic growth, evoked release, thermal protection of evoked release, and intraterminal Ca2+ levels at rest and during stimulation.
authors.affiliation: Arizona Research Laboratories Division of Neurobiology, University of Arizona, Tucson, Arizona 85721-0077, USA.
authors.forename: Peter
authors.initials: P
authors.lastname: Bronk
authors.forename: Zhiping
authors.initials: Z
authors.lastname: Nie
authors.forename: Markus K
authors.initials: MK
authors.lastname: Klose
authors.forename: Ken
authors.initials: K
authors.lastname: Dawson-Scully
authors.forename: Jinhui
authors.initials: J
authors.lastname: Zhang
authors.forename: R Meldrum
authors.initials: RM
authors.lastname: Robertson
authors.forename: Harold L
authors.initials: HL
authors.lastname: Atwood
authors.forename: Konrad E
authors.initials: KE
authors.lastname: Zinsmaier
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D030801:Animals, Genetically Modified; D005075:Biological Evolution; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology; D003952:Diagnostic Imaging / Q000379:methods; D004330:Drosophila; D029721:Drosophila Proteins / Q000378:metabolism; D015870:Gene Expression / Q000235:genetics; D050956:HSP40 Heat-Shock Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D006735:Horseradish Peroxidase / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D008564:Membrane Potentials / Q000502:physiology; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D009469:Neuromuscular Junction / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D018408:Patch-Clamp Techniques / Q000379:methods; D017354:Point Mutation / Q000502:physiology*; D017729:Presynaptic Terminals / Q000378:metabolism* / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000235:genetics / Q000502:physiology; D013329:Structure-Activity Relationship; D013696:Temperature; D013997:Time Factors
chemical_list: D029721:Drosophila Proteins; D050956:HSP40 Heat-Shock Proteins; D008565:Membrane Proteins; C088460:cysteine string protein; D006735:Horseradish Peroxidase; D002118:Calcium
references.citation: J Cell Sci. 1999 May;112 ( Pt 9):1345-51
references.pmid: 10194413
references.citation: Eur J Neurosci. 1999 May;11(5):1818-26
references.pmid: 10215934
references.citation: Curr Biol. 1999 Apr 22;9(8):R305-8
references.pmid: 10226023
references.citation: Eur J Cell Biol. 1999 Jun;78(6):375-81
references.pmid: 10430018
references.citation: Neuron. 1999 Jul;23(3):593-605
references.pmid: 10433270
references.citation: J Neurosci. 1999 Dec 1;19(23):10270-9
references.pmid: 10575024
references.citation: J Neurosci. 2000 Feb 15;20(4):1281-9
references.pmid: 10662817
references.citation: Neuron. 2000 Apr;26(1):35-43
references.pmid: 10798390
references.citation: J Neurochem. 2000 May;74(5):1781-9
references.pmid: 10800920
references.citation: J Neurosci. 2000 Aug 15;20(16):6039-47
references.pmid: 10934253
references.citation: Neuron. 2000 Oct;28(1):195-204
references.pmid: 11086994
references.citation: Biochem Pharmacol. 2001 Jul 1;62(1):1-11
references.pmid: 11377391
references.citation: Neuron. 2001 May;30(2):475-88
references.pmid: 11395008
references.citation: Neuron. 2001 Sep 27;31(6):987-99
references.pmid: 11580898
references.citation: J Physiol. 2002 Jan 15;538(Pt 2):383-9
references.pmid: 11790807
references.citation: Biochem J. 2002 Jun 1;364(Pt 2):343-7
references.pmid: 11931641
references.citation: J Comp Physiol A Neuroethol Sens Neural Behav Physiol. 2002 Feb;188(1):71-8
references.pmid: 11935231
references.citation: J Neurophysiol. 2002 Nov;88(5):2659-63
references.pmid: 12424301
references.citation: J Biol Chem. 2003 Feb 7;278(6):4258-67
references.pmid: 12458203
references.citation: J Biol Chem. 2003 Jun 6;278(23):21178-87
references.pmid: 12672808
references.citation: J Cell Sci. 2003 Jul 15;116(Pt 14):2967-74
references.pmid: 12783986
references.citation: Neuron. 1992 Sep;9(3):527-37
references.pmid: 1326297
references.citation: Nature. 1992 Apr 23;356(6371):683-9
references.pmid: 1349157
references.citation: J Biol Chem. 2003 Dec 26;278(52):53072-81
references.pmid: 14570907
references.citation: J Exp Biol. 2004 Mar;207(Pt 8):1323-34
references.pmid: 15010483
references.citation: J Neurosci. 2004 Mar 10;24(10):2496-505
references.pmid: 15014125
references.citation: Neuron. 2004 Apr 22;42(2):237-51
references.pmid: 15091340
references.citation: J Neurogenet. 1990 Nov;7(1):15-29
references.pmid: 2129171
references.citation: J Comp Neurol. 1994 Dec 15;350(3):463-72
references.pmid: 7884051
references.citation: Neuron. 1994 Sep;13(3):507-23
references.pmid: 7917288
references.citation: Neuron. 1994 Oct;13(4):899-907
references.pmid: 7946336
references.citation: J Comp Physiol A. 1994 Aug;175(2):179-91
references.pmid: 8071894
references.citation: Development. 1993 Jun;118(2):401-15
references.pmid: 8223268
references.citation: Science. 1994 Feb 18;263(5149):977-80
references.pmid: 8310297
references.citation: J Biol Chem. 1996 Oct 18;271(42):25989-93
references.pmid: 8824236
references.citation: Biochem J. 1997 Mar 15;322 ( Pt 3):853-8
references.pmid: 9148760
references.citation: Neurosci Lett. 1997 Jun 13;228(3):167-70
references.pmid: 9218634
references.citation: J Neurosci. 1997 Oct 1;17(19):7203-9
references.pmid: 9295366
references.citation: J Biol Chem. 1997 Dec 12;272(50):31420-6
references.pmid: 9395474
references.citation: J Neurosci. 1998 Feb 1;18(3):956-64
references.pmid: 9437017
references.citation: Cell. 1998 Feb 6;92(3):351-66
references.pmid: 9476895
references.citation: J Neurosci. 1998 May 1;18(9):3233-40
references.pmid: 9547232
references.citation: Nat Genet. 1998 Jun;19(2):171-4
references.pmid: 9620775
references.citation: Trends Biochem Sci. 1998 Jun;23(6):222-7
references.pmid: 9644977
references.citation: Mol Biol Cell. 1998 Aug;9(8):2259-67
references.pmid: 9693380
references.citation: EMBO J. 1998 Sep 1;17(17):5048-58
references.pmid: 9724640
references.citation: Cell Tissue Res. 1998 Nov;294(2):203-17
references.pmid: 9799436
references.citation: FEBS Lett. 1998 Oct 23;437(3):267-72
references.pmid: 9824305