Studies On The Function Of CCR5 Membrane Proximal Region In HIV-1 Infection

Since antiviral drugs,antibodies and vaccines are always in demand for the treatment or prevention of AIDS,identifying critical targets in the process of HIV-1 infection has been the focus of AIDS research.Multiple targets for antiviral drugs have been identified,including reverse transcriptase,integrase,protease,and envelop glycoprotein gp41,etc.However,these antiviral drugs always lose efficacy within a few months or weeks due to the high mutation rates of HIV-1,which is far beyond the development rates of new drugs.HIV-1 vaccines also have to face this big challenge.Herein,we focus on the conserved host protein called CCR5.We hope to find out new regions that participate in the process of HIV-1 entry and to provide novel targets for the design of HIV-1 vaccine.Potential antigenic regions on CCR5 were predicted by using bioinformatic methods.We found a region(aa 22-38)located in N-terminal domain and closely to the first transmembrane domain.This region was designated as the membrane proximal region(MPR)because of its special location.Using the C17 peptide derived from MPR as immunogen,relative high IgG titers against the C17 peptide were elicited.The MPR-specific antibodies could inhibit the infection of CCR5-tropic virus,while having no inhibition on CXCR4-tropic virus.Besides,MPR-specific antibodies neither induced the internalization of CCR5 nor inhibited the calcium influx as demonstrated by flow cytometry analysis.These results revealed that MPR played important roles in viral infection and antibodies against MPR region were more preferred than antibodies against other regions of CCR5.Other experiments were designed to validate the importance of MPR.As shown in pseudovirus infection assay,the replacement of MPR with the same region from other co-receptors significantly impaired capability of the chimeras to support viral infection.The residues I23,N24 and L32 had obvious effects on viral infection.Then the inhibitory effect of cholesterol conjugated peptide(C17-Chol)was measured by pseudovirus.C17-Chol could inhibit the infection of CCR5-tropic virus with an IC50 at low micro molar level.Furthermore,C17-Chol lost the inhibitory effect at an earlier time in viral infection than a C peptide inhibitor called C52 L.This result suggested that C17-Chol targed to an event before the formation of six helix bundle.This was coinside with the time for gp120 binding to CCR5.In conclusion,the important role of MPR in HIV-1 infection suggested that it can be a new target for antiviral drugs,antibodies or vaccines.