Archives
Arch Virol (1990) 114:265-269
Vi rology
© by Springer-Verlag 1990
Sequence homology between HIV gpl20, rabies virus glycoprotein, and snake venom neurotoxins Is the nicotinic acetylcholine receptor an HIV receptor?
Brief Report P. Neri, Luisa Bracci, M. Rustici, and Annalisa Santucci
Dipartimento di Chimica, Sezione di Chimica Medica, Universit/t di Siena, Siena, Italy Accepted June 14, 1990
Summary. We have found a striking homology between the sequence 164-174 of HIV-1 gpl20 and the sequence 30-40 of snake venom neurotoxins; this sequence homology is very similar to that existing between the same region of snake venom neurotoxins and rabies virus glycoprotein.
The nicotinic acetylcholine receptor has already been proposed as a host cell receptor for rabies virus during the infection of muscular and neuronal cells [11]. The first indication was discovered by Lentz and co-workers: they found a remarkable sequence homology between a region of the rabies virus glycoprotein and the putative functional site of snake venom curare-mimetic neurotoxins [12], which suggested a functional convergence between these different proteins. Snake venom neurotoxins are selective inhibitors of nicotinic acetylcholine receptor which compete with acetylcholine for binding to the physiologic ligand's binding sites located on receptor a-subunits [5]. Neurotoxins from Elapidae and Hydrophidae venomous snakes are small polypeptides (7,000-8,000 molecular weight); they are divided into two groups, short (60-62 residues) and long neurotoxins (71-74 residues) [reviewed in 5]; more than 60 different neurotoxins have been sequenced and comparison of their primary structures was made, accompanied by aminoacid chemical modification and studies on the three dimensional conformation: these studies provided useful information about the structure-function relations of this protein family. Snake neurotoxins share quite a constant three-dimensional structure composed of a central core and three loops: loop 2 (the toxic loop) is composed
266
P. Neri et al.
of highly conserved residues, mainly comprised in the 30--45 sequence (numbering according to [ 17]), some of which are essential for maintaining functional properties [4]. Since the homology between rabies virus glycoprotein and snake neurotoxins was first described, purified rabies virus glycoprotein was demonstrated to be able to compete with the potent neurotoxin of the snake Bungarus multicinctus a-bungarotoxin for the binding to the acetylcholine receptor I l l ; moreover anti-peptide monoclonal antibodies, directed to the glycoprotein sequence 190203 where the homology with snake neurotoxins was most evident, efficiently inhibit the binding of both rabies virus glycoprotein and a-bungarotoxin to acetylcholine receptor [1, 15], thus confirming the existence of a correlation between the snake neurotoxins functional site and a region of rabies virus glycoprotein which appears to be involved in virus binding to the receptor. While studying the correlation between rabies virus glycoprotein and snake neurotoxins we found that a similar homology with neurotoxdns toxic loop is also present in the sequence 164-174 of HIV-1 gpl20 (numbering according to [18]); comparison of HIV gpl20 with a-cobratoxin from the snake Naja ncda siarnensis shows a stretch of 5 identical residues (Fig. 1) comprising highly conserved residues among the neurotoxins family as well as the invariant R37, 038 , and K39 that are most probably involved in receptor binding (Fig. 2). Interestingly, a good homology is also evident with k-bungarotoxin, reported to be a selective marker of neuronal nicotinic receptors [7] (Fig. 1). This sequence homology may have considerable significance in view of the reported ability of HIV to infect CD4 negative cells in culture [8] and the recently described lack of inhibition by soluble CD4 [3] and anti-CD4 antibodies [19] of HIV binding to muscular and neuronal cells. In the last two cases, on the basis of their results, the authors conclude that HIV might infect neuronal and muscular cells in a way that is not mediated by CD4. The rhabdomyosarcoma cell line (TE671) used in the inhibition experiments is known to express a muscular acetylcholine receptor [16]; moreover different members of the nicotinic receptors gene family are expressed in different regions of the mammalian central nervous system [6]. I. R D
....
HRGTI
Naja nigricollis
2. S D
....
HRGTI
Dendroaspis
3. C D A F C 4. C D
S S RGKV
I F T N S RGKR
virus
5. F N I S T S I R G K V
HIV gpl20
6. C D A
~-Cobratoxin
7. C D
FC
S I R GKR
K F C S I RGPV
viridis
~-Bungarotoxin Rabies
T o x i n OC ( T x ~ ) ( 3 0 - 4 0 ) Toxin
(~-Bgt)
4.11.3(30-40)
(30-40)
glycoprotein
(189-199)
(164-174)
k-Bungarotoxin
(~-Cbt)
(30-40)
(k-Bgt)
(30-40)
Fig. 1. Comparison of aminoacid sequence of 5 HIV gpl20 (residues from 164 to 174) and 4 rabies virus glycoprotein (residues from 189 to 199) with 1, 2 short and 3, 6, 7 long neurotoxins (residues from 30 to 40)
Nicotinic receptor as an HIV receptor?
267
~5 --~GLY44
EU43
VAL41
:~P29
ARG40
'CYS30 GLY38*
~RG37@
ILE36-q---
a
b Fig. 2. a Computer graphic representation of the main chain folding of the long neurotoxin a-cobratoxin at 0.28 nm resolution. The toxic loop is protruding at the bottom in this representation, b Computer graphic representation of the toxic loop of a-cobratoxin. All the residues comprised in this loop are highly conserved among different neurotoxins except for those indicated by arrows; invariant residues are indicated by asterisks
HIV infection of muscular and neuronal cells in vitro might be mediated by gpl20 binding to nicotinic receptors present in these cell lines; the possibility that the same mechanism may be the basis of HIV neurotropism should be considered. It is k n o w n that neurologic dysfunctions occur in at least 60% of A I D S patients [10]; subacute encephalitis (AIDS encephalopathy or dementia complex) is the most c o m m o n neurologic problem which seems to be specifically induced by HIV [10, 14]. Moreover the virus has been isolated in the brain, peripheral nerves, and cerebrospinal fluid of A I D S patients with subacute encephalitis [9, 13].
268
P. Neri et al.
A sequence homology between a virus and a cellular receptor ligand, such as that observed between rabies virus or HIV and snake neurotoxins, may also be important in view o f the possibility that antibodies to the cell receptor could be produced via an anti-idiotypic immune response following an infection with the virus or, through immunization with viral proteins containing the ligandmimicking sequence. In the case of rabies virus, mice immunized with purified rabies virus glycoprotein produced auto-antibodies directed to the acetylcholine receptor [2], lost weight and died in a short time.
Acknowledgement This work was supported by a grant from CNR PF Chimica Fine.
References 1. Bracci L, Antoni G, Cusi MG, Lozzi L, Niccolai N, Petreni S, Rustici M, Santucci A, Soldani P, Valensin PE, Neff P (1988) Antipeptide monoclonal antibodies inhibit the binding of rabies virus glycoprotein and alpha-bungaratoxin to the nicotinic acetylcholine receptor. Mol Immunol 25:881-888 2. Burrage TG, Tignor GH, Smith AL (1985) Rabies virus binding at neuromuscular junctions. Virus Res 2:273-289 3. Clapham PR, Weber JN, Whitby D, McIntosh K, Dalgleish AG, Maddon PJ, Deen KC, Sweet RW, Weiss RA (1989) Soluble CD4 blocks the infectivity of diverse strains of HIV and SIV for T cells and monocytes but not for brain and muscle cells. Nature 337:368-370 4. Dufton MJ, Hider RC (1983) Conformational properties of the neurotoxins and cytotoxins isolated from elapid snake venoms. CRC Crit Rev Biochem 14:113-171 5. Endo T, Tamiya N (1987) Current view on the structure-function relationship of postsynaptic neurotoxins from snake venoms. Pharmacol Ther 34:403~451 6. Goldman D, Deneris E, Luyten W, Kochhar A, Patrick J, Heinemann S (1987) Members of the nicotinic acetylcholine receptor gene family are expressed in different regions of the mammalian central nervous system. Cell 48:965-973 7. Grant GA, Chiappinelli VA (1985) k-Bungarotoxin: complete amino acid sequence of a neuronal nicotinic receptor probe. Biochemistry 24:1532-1537 8. Harouse JM, Kunsch C, Hartle HT, Laughlin MA, Hoxie JA, Wigdahl B, GonzalesScarano F (1989) CD4-independent infection of human nem'al cells by human immunodeficiency virus type 1. J Virol 63: 2527-2533 9. Ho DD, Rota TR, Shooley RT (t985) Isolation of HTLV III from cerebrospinal fluid and neural tissues of patients with neurologic syndromes related to the acquired immunodeficiency syndrome. N Engl J Med 3t3:1493-1497 10. Ho DD, Pomerantz RJ, Kaplam JC (1987) Pathogenesis of infection with human immunodeficiency virus. N Engl J Med 317:278-286 11. Lentz TL, Burrage TG, Smith AL, Crick J, Tignor GH (1982) Is the acetylcholine receptor a rabies virus receptor? Science 215:182-184 12. Lentz TL, Wilson PT, Hawrot E, Speicher DW (1984) Amino acid sequence similarity between rabies virus glycoprotein and snake venom curaremimetic neurotoxins. Science 226:847-848 13. Levy JA, Shimabukuro J, Hollander H, Mills J, Kaminsky L (1985) Isolation of AIDSassociated retroviruses from cerebrospinal fluid and brain of patients with neurological symptoms. Lancet 2:586-588
Nicotinic receptor as an HIV receptor?
269
14. Navia BA, Cho E-S, Petito CK, Price RW (1986) The AIDS dementia complex, II: neuropathology. Ann Neurol 23:38-48 15. Rustici M, Santucci A, Lozzi L, Petreni S, Spreafico A, Neri A, Bracci L, Soldani P (1989) A monoclonal antibody to a synthetic fragment of rabies virus glycoprotein binds ligands of the nicotinic cholinergic receptor. J Mol Rec 2:51-55 16. Schoepfer R, Luther M, Lindstrom J (1988) The human medulloblastoma cell line TE671 expresses a muscle-like acetylcholine receptor. FEBS Lett 226:235--240 17. Schwartz RM, Dayhoff MO (1978) Atlas of protein sequence and structure, vo15, suppl 3. National Biomedical Research Foundation, Washington, DC 18. Wain-Hobson S, Sonigo P, Danos O, Cole S, Alizon M (1985) Nucleotide sequence of the AIDS virus, LAV. Cell 40:9-17 19. Weber J, Clapham P, McKeating J, Stratton M, Robey E, Weiss R (1989) Infection of brain cells by diverse Human ImmunodeficiencyVirus isolates: role of CD4 receptor. J Gen Virol 70:2653-2660 Authors' address: Luisa Bracci, Dipartimento di Chimica, Sezione di Chimica Medica, Universitfi di Siena, Policlinico Le Scotte, 1-53100 Siena, Italy. Received June 14, 1990
Arch Virol (1990) 114:265-269
Vi rology
© by Springer-Verlag 1990
Sequence homology between HIV gpl20, rabies virus glycoprotein, and snake venom neurotoxins Is the nicotinic acetylcholine receptor an HIV receptor?
Brief Report P. Neri, Luisa Bracci, M. Rustici, and Annalisa Santucci
Dipartimento di Chimica, Sezione di Chimica Medica, Universit/t di Siena, Siena, Italy Accepted June 14, 1990
Summary. We have found a striking homology between the sequence 164-174 of HIV-1 gpl20 and the sequence 30-40 of snake venom neurotoxins; this sequence homology is very similar to that existing between the same region of snake venom neurotoxins and rabies virus glycoprotein.
The nicotinic acetylcholine receptor has already been proposed as a host cell receptor for rabies virus during the infection of muscular and neuronal cells [11]. The first indication was discovered by Lentz and co-workers: they found a remarkable sequence homology between a region of the rabies virus glycoprotein and the putative functional site of snake venom curare-mimetic neurotoxins [12], which suggested a functional convergence between these different proteins. Snake venom neurotoxins are selective inhibitors of nicotinic acetylcholine receptor which compete with acetylcholine for binding to the physiologic ligand's binding sites located on receptor a-subunits [5]. Neurotoxins from Elapidae and Hydrophidae venomous snakes are small polypeptides (7,000-8,000 molecular weight); they are divided into two groups, short (60-62 residues) and long neurotoxins (71-74 residues) [reviewed in 5]; more than 60 different neurotoxins have been sequenced and comparison of their primary structures was made, accompanied by aminoacid chemical modification and studies on the three dimensional conformation: these studies provided useful information about the structure-function relations of this protein family. Snake neurotoxins share quite a constant three-dimensional structure composed of a central core and three loops: loop 2 (the toxic loop) is composed
266
P. Neri et al.
of highly conserved residues, mainly comprised in the 30--45 sequence (numbering according to [ 17]), some of which are essential for maintaining functional properties [4]. Since the homology between rabies virus glycoprotein and snake neurotoxins was first described, purified rabies virus glycoprotein was demonstrated to be able to compete with the potent neurotoxin of the snake Bungarus multicinctus a-bungarotoxin for the binding to the acetylcholine receptor I l l ; moreover anti-peptide monoclonal antibodies, directed to the glycoprotein sequence 190203 where the homology with snake neurotoxins was most evident, efficiently inhibit the binding of both rabies virus glycoprotein and a-bungarotoxin to acetylcholine receptor [1, 15], thus confirming the existence of a correlation between the snake neurotoxins functional site and a region of rabies virus glycoprotein which appears to be involved in virus binding to the receptor. While studying the correlation between rabies virus glycoprotein and snake neurotoxins we found that a similar homology with neurotoxdns toxic loop is also present in the sequence 164-174 of HIV-1 gpl20 (numbering according to [18]); comparison of HIV gpl20 with a-cobratoxin from the snake Naja ncda siarnensis shows a stretch of 5 identical residues (Fig. 1) comprising highly conserved residues among the neurotoxins family as well as the invariant R37, 038 , and K39 that are most probably involved in receptor binding (Fig. 2). Interestingly, a good homology is also evident with k-bungarotoxin, reported to be a selective marker of neuronal nicotinic receptors [7] (Fig. 1). This sequence homology may have considerable significance in view of the reported ability of HIV to infect CD4 negative cells in culture [8] and the recently described lack of inhibition by soluble CD4 [3] and anti-CD4 antibodies [19] of HIV binding to muscular and neuronal cells. In the last two cases, on the basis of their results, the authors conclude that HIV might infect neuronal and muscular cells in a way that is not mediated by CD4. The rhabdomyosarcoma cell line (TE671) used in the inhibition experiments is known to express a muscular acetylcholine receptor [16]; moreover different members of the nicotinic receptors gene family are expressed in different regions of the mammalian central nervous system [6]. I. R D
....
HRGTI
Naja nigricollis
2. S D
....
HRGTI
Dendroaspis
3. C D A F C 4. C D
S S RGKV
I F T N S RGKR
virus
5. F N I S T S I R G K V
HIV gpl20
6. C D A
~-Cobratoxin
7. C D
FC
S I R GKR
K F C S I RGPV
viridis
~-Bungarotoxin Rabies
T o x i n OC ( T x ~ ) ( 3 0 - 4 0 ) Toxin
(~-Bgt)
4.11.3(30-40)
(30-40)
glycoprotein
(189-199)
(164-174)
k-Bungarotoxin
(~-Cbt)
(30-40)
(k-Bgt)
(30-40)
Fig. 1. Comparison of aminoacid sequence of 5 HIV gpl20 (residues from 164 to 174) and 4 rabies virus glycoprotein (residues from 189 to 199) with 1, 2 short and 3, 6, 7 long neurotoxins (residues from 30 to 40)
Nicotinic receptor as an HIV receptor?
267
~5 --~GLY44
EU43
VAL41
:~P29
ARG40
'CYS30 GLY38*
~RG37@
ILE36-q---
a
b Fig. 2. a Computer graphic representation of the main chain folding of the long neurotoxin a-cobratoxin at 0.28 nm resolution. The toxic loop is protruding at the bottom in this representation, b Computer graphic representation of the toxic loop of a-cobratoxin. All the residues comprised in this loop are highly conserved among different neurotoxins except for those indicated by arrows; invariant residues are indicated by asterisks
HIV infection of muscular and neuronal cells in vitro might be mediated by gpl20 binding to nicotinic receptors present in these cell lines; the possibility that the same mechanism may be the basis of HIV neurotropism should be considered. It is k n o w n that neurologic dysfunctions occur in at least 60% of A I D S patients [10]; subacute encephalitis (AIDS encephalopathy or dementia complex) is the most c o m m o n neurologic problem which seems to be specifically induced by HIV [10, 14]. Moreover the virus has been isolated in the brain, peripheral nerves, and cerebrospinal fluid of A I D S patients with subacute encephalitis [9, 13].
268
P. Neri et al.
A sequence homology between a virus and a cellular receptor ligand, such as that observed between rabies virus or HIV and snake neurotoxins, may also be important in view o f the possibility that antibodies to the cell receptor could be produced via an anti-idiotypic immune response following an infection with the virus or, through immunization with viral proteins containing the ligandmimicking sequence. In the case of rabies virus, mice immunized with purified rabies virus glycoprotein produced auto-antibodies directed to the acetylcholine receptor [2], lost weight and died in a short time.
Acknowledgement This work was supported by a grant from CNR PF Chimica Fine.
References 1. Bracci L, Antoni G, Cusi MG, Lozzi L, Niccolai N, Petreni S, Rustici M, Santucci A, Soldani P, Valensin PE, Neff P (1988) Antipeptide monoclonal antibodies inhibit the binding of rabies virus glycoprotein and alpha-bungaratoxin to the nicotinic acetylcholine receptor. Mol Immunol 25:881-888 2. Burrage TG, Tignor GH, Smith AL (1985) Rabies virus binding at neuromuscular junctions. Virus Res 2:273-289 3. Clapham PR, Weber JN, Whitby D, McIntosh K, Dalgleish AG, Maddon PJ, Deen KC, Sweet RW, Weiss RA (1989) Soluble CD4 blocks the infectivity of diverse strains of HIV and SIV for T cells and monocytes but not for brain and muscle cells. Nature 337:368-370 4. Dufton MJ, Hider RC (1983) Conformational properties of the neurotoxins and cytotoxins isolated from elapid snake venoms. CRC Crit Rev Biochem 14:113-171 5. Endo T, Tamiya N (1987) Current view on the structure-function relationship of postsynaptic neurotoxins from snake venoms. Pharmacol Ther 34:403~451 6. Goldman D, Deneris E, Luyten W, Kochhar A, Patrick J, Heinemann S (1987) Members of the nicotinic acetylcholine receptor gene family are expressed in different regions of the mammalian central nervous system. Cell 48:965-973 7. Grant GA, Chiappinelli VA (1985) k-Bungarotoxin: complete amino acid sequence of a neuronal nicotinic receptor probe. Biochemistry 24:1532-1537 8. Harouse JM, Kunsch C, Hartle HT, Laughlin MA, Hoxie JA, Wigdahl B, GonzalesScarano F (1989) CD4-independent infection of human nem'al cells by human immunodeficiency virus type 1. J Virol 63: 2527-2533 9. Ho DD, Rota TR, Shooley RT (t985) Isolation of HTLV III from cerebrospinal fluid and neural tissues of patients with neurologic syndromes related to the acquired immunodeficiency syndrome. N Engl J Med 3t3:1493-1497 10. Ho DD, Pomerantz RJ, Kaplam JC (1987) Pathogenesis of infection with human immunodeficiency virus. N Engl J Med 317:278-286 11. Lentz TL, Burrage TG, Smith AL, Crick J, Tignor GH (1982) Is the acetylcholine receptor a rabies virus receptor? Science 215:182-184 12. Lentz TL, Wilson PT, Hawrot E, Speicher DW (1984) Amino acid sequence similarity between rabies virus glycoprotein and snake venom curaremimetic neurotoxins. Science 226:847-848 13. Levy JA, Shimabukuro J, Hollander H, Mills J, Kaminsky L (1985) Isolation of AIDSassociated retroviruses from cerebrospinal fluid and brain of patients with neurological symptoms. Lancet 2:586-588
Nicotinic receptor as an HIV receptor?
269
14. Navia BA, Cho E-S, Petito CK, Price RW (1986) The AIDS dementia complex, II: neuropathology. Ann Neurol 23:38-48 15. Rustici M, Santucci A, Lozzi L, Petreni S, Spreafico A, Neri A, Bracci L, Soldani P (1989) A monoclonal antibody to a synthetic fragment of rabies virus glycoprotein binds ligands of the nicotinic cholinergic receptor. J Mol Rec 2:51-55 16. Schoepfer R, Luther M, Lindstrom J (1988) The human medulloblastoma cell line TE671 expresses a muscle-like acetylcholine receptor. FEBS Lett 226:235--240 17. Schwartz RM, Dayhoff MO (1978) Atlas of protein sequence and structure, vo15, suppl 3. National Biomedical Research Foundation, Washington, DC 18. Wain-Hobson S, Sonigo P, Danos O, Cole S, Alizon M (1985) Nucleotide sequence of the AIDS virus, LAV. Cell 40:9-17 19. Weber J, Clapham P, McKeating J, Stratton M, Robey E, Weiss R (1989) Infection of brain cells by diverse Human ImmunodeficiencyVirus isolates: role of CD4 receptor. J Gen Virol 70:2653-2660 Authors' address: Luisa Bracci, Dipartimento di Chimica, Sezione di Chimica Medica, Universitfi di Siena, Policlinico Le Scotte, 1-53100 Siena, Italy. Received June 14, 1990
