Amphotropic Murine Leukemia Virus Entry Is Determined by Specific Combinations of Residues from Receptor Loops 2 and 4

Abstract

Pit2 is the human receptor for amphotropic murine leukemia virus (A-MuLV); the related human protein Pit1 does not support A-MuLV entry. Interestingly, chimeric proteins in which either the N-terminal or the C-terminal part of Pit2 was replaced by the Pit1 sequence all retained A-MuLV receptor function. A possible interpretation of these observations is that Pit1 harbors sequences which can specify A-MuLV receptor function when presented in a protein context other than Pit1, e.g., in Pit1-Pit2 hybrids. We reasoned that such Pit1 sequences might be identified if presented in the Neurospora crassa protein Pho-4. This protein is distantly related to Pit1 and Pit2, predicted to have a similar membrane topology with five extracellular loops, and does not support A-MuLV entry. We show here that introduction of the Pit1-specific loop 2 sequence conferred A-MuLV receptor function upon Pho-4. Therefore, we conclude that (i) a functional A-MuLV receptor can be constructed by combining sequences from two proteins each lacking A-MuLV receptor function and that (ii) a Pit1 sequence can specify A-MuLV receptor function when presented in another protein context than that provided by Pit1 itself. Previous results indicated a role of loop 4 residues in A-MuLV entry, and the presence of a Pit2-specific loop 4 sequence was found here to confer A-MuLV receptor function upon Pho-4. Moreover, the introduction of a Pit1-specific loop 4 sequence, but not of a Pit2-specific loop 4 sequence, abolished the A-MuLV receptor function of a Pho-4 chimera harboring the Pit1-specific loop 2 sequence. Together, these data suggest that residues in both loop 2 and loop 4 play a role in A-MuLV receptor function. A-MuLV is, however, not dependent on the specific Pit2 loop 2 and Pit2 loop 4 sequences for entry; rather, the role played by loops 2 and 4 in A-MuLV entry can be fulfilled by several different combinations of loop 2 and loop 4 sequences. We predict that the residues in loops 2 and 4, identified in this study as specifying A-MuLV receptor function, are to be found among those not conserved among Pho-4, Pit1, and Pit2.

Mapping of Equine Lentivirus Receptor 1 Residues Critical for Equine Infectious Anemia Virus Envelope Binding

“…The observation that no single point mutation in ELR1 completely abrogated gp90 binding suggests further that receptor binding to the viral envelope protein likely involves multiple contact points with residues within and outside of the CRD1 sequences. Thus, these data are consistent with other reports that multiple amino acid point mutations are required to completely eliminate receptor activity for FIV (11), avian sarcomaleukosis virus (16,17), human immunodeficiency virus (23), and murine leukemia virus (21).…”

Evolutionary and experimental analyses of inorganic phosphate transporter PiT family reveals two related signature sequences harboring highly conserved aspartic acids critical for sodium‐dependent phosphate transport function of human PiT2

“…Moreover, the hypothesis that region A should be intracellular in PiT1 and extracellular in PiT2 seems in conflict with the observation that a PiT2 mutant with PiT1 region A sequence is receptor for both PiT1 and PiT2 cognate viruses [36] but shows unimpaired P i uptake function compared to PiT2 (P. Bøttger and L. Pedersen, unpublished work). Taken together the current data on receptor and transport functions of PiT1, PiT2, Pho4, and chimeras derived from these are all in favor [23,26,36,37,[45][46][47][48][49][50][51][52][53][54][55], or at least are not in conflict with [41], an extracellular position of region A. Thus, the most likely membrane topology of both PiT1 and PiT2 is that proposed by Salau¨n et al for PiT2 [23] (Fig.…”

Use of different but overlapping determinants in a retrovirus receptor accounts for non-reciprocal interference between xenotropic and polytropic murine leukemia viruses

“…Initial predictions of receptor topology were used to design a number of chimeras similar to those described here. Regions within those chimeras were identified that enhanced infection by GALV or amphotropic MLV respectively, and it was suggested that these regions were responsible for virus binding [ 30 - 33 ]. However, recent experiments have provided a new, experimentally verified topology for Pit2 [ 34 ], and several of the previously identified critical regions were found to lie on the inner surface of the cell membrane.…”