A cohort-based study of host gene expression: tumor suppressor and innate immune/inflammatory pathways associated with the HIV reservoir size

The major barrier to an HIV cure is the HIV reservoir: latently-infected cells that persist despite effective antiretroviral therapy (ART). Most prior studies of host genetic predictors of HIV control have focused on “elite controllers,” rare individuals able to control virus in the absence of ART. However, there have been few genetic studies among ART-suppressed non-controllers, who make up the majority of people living with HIV (PLWH). We performed host RNA sequencing and HIV reservoir quantification (total DNA [tDNA], unspliced RNA [usRNA], intact DNA) from peripheral CD4+ T cells from 191 HIV+ ART-suppressed non-controllers. After adjusting for nadir CD4+ count, timing of ART initiation, and genetic ancestry, we identified two host genes for which higher expression was significantly associated with smaller total DNA viral reservoir size, P3H3 and NBL1, both known tumor suppressor genes. We then identified 17 host genes for which lower expression was associated with higher residual t..., HIV+ ART-suppressed non-controllers from the UCSF SCOPE and Options HIV+ cohorts were included in the study.
Cryopreserved PBMCs were enriched for CD4+ T cells (StemCell, Vancouver, Canada), and RNA was extracted from CD4+ T cells using the AllPrep Universal Kit (Qiagen, Hilden, Germany) with one aliquot set aside for HIV reservoir quantification and a second aliquot for host RNA sequencing. Host RNA sequencing was analyzed using the HTStream pre-processing pipeline (s4hts.github.io/htstream/) was used for removing PCR duplicates, adapters, N characters, PolyA/T sequences, Phix contaminants, and poor-quality sequences (with quality score <20 with sliding window of 10 base pairs). The quality of raw reads was assessed using FastQC. All samples had a per base quality score and sequence quality score >30. RNA-seq reads were then mapped to the human genome (GRCh38) with a corresponding annotation file from the GENCODE project. Alignment and gene quantification were performed using the..., , # A cohort-based study of host gene expression: tumor suppressor and innate immune/inflammatory pathways associated with the HIV reservoir size

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These data include host bulk RNA-seq data from peripheral CD4+ T cells from 191 people with HIV-infected adults, virally suppressed on antiretroviral therapy (ART). Cryopreserved PBMCs were enriched for CD4+ T cells (StemCell, Vancouver, Canada), and RNA was extracted from CD4+ T cells using the AllPrep Universal Kit (Qiagen, Hilden, Germany). For validation of intracellularly expressed or membrane-associated encoded proteins, we performed ELISA in a subset of the participants, using matched peripheral CD4-enriched T cells of 5 proteins (Kir2.1, connexin 26, P3H3, NBL1 and TLR4) encoded by KCNJ2, GJB2, P3H3, NBL1 and TLR4 ...

HIV治愈研究的核心障碍在于HIV储存库（HIV reservoir）：即即便接受有效的抗逆转录病毒治疗（antiretroviral therapy, ART）仍可持续存在的潜伏感染细胞。既往绝大多数关于HIV宿主遗传调控因素的研究，均聚焦于"精英控制者"——这类罕见个体无需ART即可自主抑制病毒复制。然而，针对接受ART病毒抑制治疗的非控制者群体的遗传研究却极为匮乏，而该群体占全球艾滋病病毒感染者（people living with HIV, PLWH）的绝大多数。

本研究针对191名接受ART抑制治疗的HIV阳性非控制者，对其外周CD4+ T细胞（CD4-positive T cells）开展了宿主RNA测序（RNA-seq）及HIV储存库定量检测（涵盖总DNA[tDNA]、未剪接RNA[usRNA]、完整DNA三个指标）。在校正了最低CD4+计数、ART启动时机及遗传血统等混杂变量后，我们鉴定出两个宿主基因：其表达水平越高，病毒总DNA储存库规模越小，即P3H3与NBL1，二者均为已报道的抑癌基因。随后我们进一步鉴定出17个宿主基因，其表达水平越低则残留tDNA水平越高[原文此处存在截断]。本研究纳入了来自加州大学旧金山分校（UCSF）SCOPE队列与Options HIV队列的接受ART抑制治疗的HIV阳性非控制者。

我们对冻存的外周血单个核细胞（peripheral blood mononuclear cell, PBMC）进行CD4+ T细胞富集（使用加拿大温哥华StemCell公司的相关试剂），并采用德国希尔德市Qiagen公司的AllPrep Universal试剂盒从CD4+ T细胞中提取RNA：将样本分为两等份，一份用于HIV储存库定量检测，另一份用于宿主RNA测序。宿主RNA测序数据采用HTStream预处理流程（https://s4hts.github.io/htstream/）进行分析，该流程可去除PCR重复序列、测序接头、N碱基、PolyA/T均聚物序列、PhiX噬菌体污染物及低质量序列（采用10bp滑动窗口，过滤质量分数<20的序列）。原始测序读段的质量采用FastQC进行评估，所有样本的每碱基质量分数及整体序列质量分数均高于30。随后，我们将RNA-seq读段比对至人类参考基因组GRCh38，并使用GENCODE项目提供的对应注释文件。比对分析与基因定量工作采用[原文此处存在截断]。

# 基于队列的宿主基因表达研究：与HIV储存库规模相关的抑癌基因及先天免疫/炎症通路

本数据集包含191名接受抗逆转录病毒治疗且病毒得到抑制的HIV感染成人的外周CD4+ T细胞宿主批量RNA测序（bulk RNA-seq）数据。我们对冻存的PBMC进行CD4+ T细胞富集（使用加拿大温哥华StemCell公司的相关试剂），并采用德国希尔德市Qiagen公司的AllPrep Universal试剂盒从CD4+ T细胞中提取RNA。为验证细胞内表达或膜结合的编码蛋白，我们在部分参与者中开展了酶联免疫吸附试验（enzyme-linked immunosorbent assay, ELISA），针对由KCNJ2、GJB2、P3H3、NBL1及TLR4基因编码的5种蛋白（Kir2.1、连接蛋白26、P3H3、NBL1及TLR4）进行检测。

本数据集的官方DOI链接为：https://doi.org/10.5061/dryad.k3j9kd5dw