APOBEC3G Variant (rs6001417) CG and GG Genotypes and their protective feature against HIV-1 Infection in Pakistani Dwelled Community

Full Length Research Article

APOBEC3G Variant (rs6001417) CG and GG Genotypes and their protective feature against HIV-1 Infection in Pakistani Dwelled Community

Qaisar Ali2, Arshad Jamal1,2 , Sajjad Ullah2, Ahmed Bilal Waqar*2

Adv. life sci., vol. 8, no. 2, pp. 108-113, February 2021
*Corresponding Author: Ahmed Bilal Waqar (Email: drabwaqar@yahoo.com)
Authors' Affiliations

 1. Department of Biology, College of Science. University of Hail – Kingdom of Saudi Arabia
2. Department of Medical Laboratory Sciences (DMLS), Faculty of Allied and Health Sciences, Imperial College of Business Studies Lahore – Pakistan
 [Date Received: 10/08/2020; Date Revised: 08/10/2020; Date Published: 25/02/2021]

Abstractaa download_button



Background: APOBEC3G (Apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like 3G) gene is one of the genetic host factors, have been linked with HIV-1 AIDS predisposing and protection in different residence populations. The investigation of genetic marker (APOBEC3G) variant (rs6001417) CC, CG and GG genotypes in Pakistan.

Methods: The extraction of DNA, the DNA Rapid Salting-out method was used. Then the observed DNA with electrophoresis technique referred for quantitative real-time PCR to identify the APOBEC3G variant rs6001417 genotypes and Taq Man genotyping.  

Results: Three genotypes of rs6001417 (CC, CG and GG) were compared both in HIV-1 infected patients and healthy control groups (p=0.73, p=0.007, p=0.01 respectively). The rs6001417 CG and GG genotype demonstrated a significant involvement in both the healthy and infected individuals and portraying possible protective effect against HIV-1 infection with predictive value of 36.43% and 13.57% respectively.

Conclusion: APOBEC3G (rs6001417) CG and GG genotypes may have a protective feature in the progression of HIV-1 infection and we may use this as a preliminary predictive marker in the country for HIV-1 infected individuals as well.

Keywords: HIV-1; APOBEC3G; Predictive marker; Predictive value; Real-time PCR

Introduction6th button-01

HIV-1 infection is one of the global health problems, affecting almost 37.9 as million human population as of 2019 according to UNAIDS worldwide, with 3.1 million new cases  reported every year [1-3, 40]. In recent past, HIV infection is highly prevalent in Sub-Saharan countries [4], however in Pakistan, currently it is one of the leading causes of morbidity and mortality as it hits approximately 160,000 infected individuals according to UN. Mostly HIV infection is being transmitted via contaminated blood and sexual contacts [5,6].

Immune related genetic mechanism can participate with the immune system while excluding a specific type of antigen from the body [7-11]. Recent findings portray that, both the host and viral genetics may have a substantial influence in the disease progression and protection [12-14]. In case of homozygous allelic variant of CCR5 protein has a substantial contribution in against the HIV infection [15,16]. Also, the ethnic background and DNA sequence similarity has a great role in the susceptibility and protection of a disease [17,19]. The only reason attracts  researchers globally to study the genetic role in the progression of HIV infection.

In recent times, several studies have demonstrated that multiple host factors have influenced the pathogenesis of HIV-1/AIDS condition. These factors includes; APOBEC3G, Chemokine Receptor 5 (CCR-5), Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Non-integrin (DC-SIGN), Tripartite motif 5a (TRIM5a), Tetherin, and (SAM-domain HD-domain containing protein) SAMHD1 [5,20-23]. These are antagonized by accessory viral proteins [24,25].

Moreover, Apolipoprotein B mRNA editing enzyme catalytic polypeptide-like 3G (APOBEC3G) is an effective factor inside the host, which interferes with  HIV-1 [22]. Virion infectivity factor (vif) which is structural part of  HIV-1 is able to counteract APOBEC3G antiviral activity by targeting it for degradation in proteasomes [26,27]. Vif proteins derived from subtypes A, B, CRF01_AE, and CRF_02AG  showed non-significant but some-what differential anti-APOBEC3G activity levels based on infectivity profiles while subtype C was highly significant [28,29]. The APOBEC3G protein was incorporated into newly synthesized viral particles, in the absence of the virion infectivity factor (vif), and deamination of cytosine (C) to uracil (U) made viral DNA mutated. APOBEC3G polymorphisms, such as (H186R) rs8177832, are supposed to be related with HIV-1 subtype B and C pathogenesis in different ethnic groups [7,30], however this association is not  found in other populations [31-33]. These previous studies did not take the Circulating Recombinant Forms of HIV-1 into consideration, nor examine the effect of APOBEC3G polymorphisms in Asian Pakistani ethnic groups. The present study was  conducted to understand and make clear the role of rs6001417, variants of APOBEC3G in HIV-1 infection in Pakistani  population.

Methods6th button-01

Study population

A total of 240 subjects (100 patients and 140 healthy persons) were included in this study.  Samples were collected from different HIV centers of Pakistan and processed at Imperial Diagnostics and Research Center, Lahore.

Sample collection, HIV-1 testing  

Intravenous blood  samples collected from HIV-1 infected subjects were genotyped using Real Time PCR Quantitative kit (SYBR GreenER) 100-rxn  according to manufacturer’s instructions.

DNA extraction and genotyping

The DNA  was  extracted from blood  by using the “DNA Rapid Salting-out” procedure which has characterized by Miller et al [34].  The  extracted DNA  was stored at -20oC till  further processing. The  concept of SNP  was based on A3G variant rs6001417 along with the defense in contact with HIV-1  infection  [30]. The rs6001417 in regard to A3G  was genotyped by applying common SYBR GreenER SNP assay on the Fast Real-Time PCR Systems (Applied Biosystem Step One TM). PCR amplification was performed using the reverse and  forward primers  respectively Table 1. Every reaction was consisted of absolute amount of 25 µl, containing 10x PCR buffer 2.5 µl, Taq polymerase 0.5 µl, d-NTPs 1 µl,  each primer 1 µl, genomic DNA 1 µl. The processing started by denaturation at 95℃, 30s of annealing at 55oC and 30s of extension  at 68℃, in 35 cycles. The final extension was, at 72℃ for 7 minutes. After electrophoresis over a 2 percent agarose gel with 0.5 ug/ml ethidium bromide, the amplified product was  examined on  UV light.

Ethical considerations

The study  was approved from the  Institutional Ethical Committee of Imperial College of Business Studies and written consent from each participant was obtained.

Statistical analysis

The epidemiological data were recorded on a pre-designed form and handled within excel software. All calculations were  performed by applying SPSS software version 20.0 statistical package. Data is expressed as Mean ± S.D and calibrations like diagnostic aspects has been calculated. Categorical variable was analyzed with X2 test. Hardy Weinberg equilibrium is also applied for Allele frequency. Contrasts to genotype placement of the groups were determined through the X2 trial. P<0.05 is considered as statistically significant.  

Results6th button-01

A total of 240 individuals were  included into the study. Study population consisted of 100 HIV-1 infected patients and 140 healthy controls. Gender wise distribution, age of the cases and healthy groups have been defined in Table 2. The mean age of the study population was 39.21±11.7 in the healthy control group and 35.94±9.84 in the HIV-1 cases group. We found gender and age comparable variables among the two groups (P< 0.05). Genotype frequencies of the one APOBEC3G loci (rs6001417) for both the cases and controls  were given correspondingly in Table 2.

1. Association between APOBEC3G variants and HIV-1 status

We  analyzed and compared SNP rs6001417 genotypes ( CC, CG and GG) between cases and control groups by using P-value of  > 0.05, as shown in Table 2. The electrophoresis pattern of APOBEC3G (rs6001417) CC, CG, and GG genotypes  was shown in Figure 1.

1.1 APOBEC3G (rs6001417)

The genotype frequencies of  APOBEC3G (rs6001417) CC, CG, and GG genotypes were 27.50%, 11.20%, 2.90% in the patient with HIV-1 group and 29.20%, 21.20%, 7.90% in the control group, respectively as shown in Table 1. Individuals account for HIV-1 infection has lower frequencies of the APOBEC3G (rs6001417) CG,GG genotypes than healthy individuals while CC   in HIV-1 patients. Chi-square analysis provided information that rs6001417 CG and GG genotypes (51 (21.20%) vs 27 (11.20%) ; p=0.007 and 19 (7.90%) vs 07 (2.90%); p=0.019) reflect a significant variation between the two groups as shown in Figure 2. This analysis showed that subjects account for rs6001417 CG and GG genotype displays a protective role toward the HIV-1 infection.

2. Association of gender with protective and predisposing attaining APOBEG3G variants genotypes

We found rs6001417 CC,CG and GG genotype frequencies and distribution as 42.10%, 20.00%, 07.10% and 14.60%, 12.50%, 3.80%  in both male and female gender, respectively and had found comparable (P< 0.05) Table 2, Figure 3.

2.1 Analysis of gender with Protective (rs6001417) CG and GG genotypes

The rs6001417 CG and GG genotype had a comparable association in both the male and female studied population (20.00% vs 12.50%; P= 0.062 and 07.10% vs 3.80%; p=0.117) (Table 2). This analysis showed that, there is a comparable contribution of both male and female towards the protection in CG and GG individuals (reduces risk of being infected) of the HIV-1 studied population. Moreover, (rs6001417) CC genotype  was more common in male than female in HIV-1 studied population and statistically significant (42.10% vs 14.60%; p <0.001) as shown in Table 2. These findings showed the maximum contribution by male gender towards the CC genotype which is being insignificant in both the HIV-1 cases and control groups.  These comparisons were noticed in the HIV-1 studied population only.

3. Predictive value of APOBEG3G (rs6001417) CG and GG genotypes

We have already discussed  the protective rs6001417 CG, and GG genotypes for HIV-1 infection. In addition, we examined predictive value of these two genotypes as well. Both the rs6001417 CG and GG genotypes were  found to be the protective genotypes, we also  calculated positive predictive value (PPV) which was 36.43% and 13.57%, respectively as shown in Table 3.


Figures & Tables












Discussion6th button-01

Continual exposure to HIV infection does not certainly result in AIDS occurrence [35]. Multiple genetic and immune factors help in HIV acquirement, pathogenesis and AIDS progression at various stages of HIV life-cycle. So, HIV infection activates multiple intrinsic host factors that confer resistance to HIV pathogenesis, though the most important one is  intrinsic inhibition to HIV infection  by APOBEC3G genetic host factor [24,36]. APOBEC3G single nucleotide polymorphisms (SNPs) are of particular importance and its twenty-nine SNPs have been studied in American [37] and European [38] cohorts to reveal its influence on AIDS development and progression. We examined the frequency distribution of the variants rs6001417 of APOBEC3G gene in the population of Pakistan. Moreover, we studied the APOBEC3G gene polymorphism without the fact of that virion infectivity factor (vif), as it degrades the HIV-1 virus along with APOBEC3G gene [26,27].

The most common genotype was CC of APOBEC3G variants (rs6001417) followed by CG, GG , in the whole studied population. Furthermore, it was observed, that APOBEC3G rs6001417 CG and GG genotypes provide significant protection with 36.43% and 13.57% positive predictive value (PPV) respectively, towards HIV-1 infection. We have already reported predictive value in the treatment of HCV infection recently in which IL28B rs12979860 CT predicted 81.56% [39]. In a recently reported study from Pakistan, has shown that rs8177832 AA genotype has a predisposing role whereas, rs8177832 AG genotype portrayed a protective prediction against HIV-1 disease [40]. Recently a study conducted in Burkina Faso revealed that APOBEC3G GGT haplotypes for rs6001417 variants have an influential outcome by providing protection against HIV infection in comparison to other haplotypes. The  results of the same study also demonstrated that individuals with haplotypes GGC  has an increase of two- to five-folds in susceptiblity against the HIV  infecton [41]. However, French cohort study showed a comparable association of APOBEC3G genetic variation, H186R, with the disease progression [31]. Interestingly, Tegwinde Rebeca Compaore et.al reported that rs6001417 GG and CG genotypes predicted protection and predisposing factor against the HIV-1 respectively, which is in line with our study that also showed both rs6001417 GG and CG genotypes with protective feature [42].

Moreover, in the recent past APOBEC3G was also studied with HBV infection, though a Moroccan based study where 179 chronic infected inviduals along with 216 control were involved concluded with comparable results after testing hypothesis [43]. But, this variation of the results might be due to the distinct level of population and genetic makeup. This gentic variation was noted as higher as 37%, 3% and 5% in African Americans, Europian Americans and europian, respectively [7]. Similarly, Single Nucleotide polymorphisms of APOBEC3G docking proteins such as Vif and CUL5 can also help in the progression of the disease [44].

According to our studied population, the effect of APOBEC3G rs6001417 CC (42.10% vs 14.60% ; P = <0.001) genotype with in gender as it  found frequently in males compare to female. Therefore, the notion that the rs6001417 CT genotype may be supported by the male in term of protection or predisposing against HIV-1 infection is not supported by our results because we found this CT genotype insignificant (neither protective nor predisposing). Interestingly,we reported gender association in recent past with IL28B rs12979860 CT genotype  against the spotaneous clearance of HCV infection, in which female gender supported CT genotype in term of spontaneous clearance of HCV infection [45].  To the best of our knowledge, no one has studied or reported the gender interaction with APOBEC3G gene polymorphism and HIV-1 infection. The limitations of this interaction, need further investigations as the sample size of our study was not enough and need further investigation in order to clarify the association.

APOBEC3G polymorphisms variants (rs6001417) CG and GG genotypes may play a vital role at biological level in the interaction of HIV-1 susceptibility to the host, however, extra efforts are required on a larger cohort of patients to elucidate the association.

Author Contributions

All authors contributed equally in preparation and publication of this manuscript.

Conflict of Interest

The authors declare that there is no conflict of interest regarding the publication of this paper.


This work was supported by Çanakkale Onsekiz Mart University the Scientific Research Coordination Unit, Project number: 2008-44.

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