Interactive association of heat shock protein 70 gene variants with natural longevity in Xinjiang Hetian Uygur ethnicity.

JINXIN LI, WENQUAN NIU, YUE QI, WUFUER MAYILA, PENGCHENG ZHU, MUHUYATI, ZUHENG CHENG, and CHANGCHUN QIU from the Cardiology Department, The First Affiliated Hospital of Xinjiang Medical University, Urumqi, Xinjiang, China; State Key Laboratory of Medical Genomics; Sino-French Research Center for Life Science and Genomics, Ruijin Hospital, Shanghai Institute of Hypertension, Shanghai Jiaotong University School of Medicine, Shanghai, China; Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China. 

Mounting evidence suggests that all organisms at the cellular level respond to stress by synthesizing heat shock proteins at the expense of other proteins. The ability of human cells to respond to heat stress decreases with aging. We thus investigate the association of 3 variants (A1267G in HSPA1B, G190C in HSPA1A, and T2437C in HSPA1L) in the heat shock protein 70 (Hsp70) family with natural longevity in a Xinjiang Hetian Uygur population. A case-control study was conducted in 191 healthy individuals greater than 90 years of age and 53 naturally died persons 65-70 years of age. Promoter activity was evaluated by luciferase reporter assays. The data were analyzed using an EH/EH1 program for haplotype prediction and MDR software for gene-gene interaction. All studied variants satisfied the Hardy-Weinberg equilibrium in each group.

In single-locus analysis, no significant differences were found between long-lived people and short-lived people in the genotype/allele distributions of all variants. In contrast, haplotype analysis indicated that haplotypes A-G-C and A-C-T were more prevalent in long-lived people than short-lived people (P=0.026 and 0.017). The analysis conferred a 3.46- and 4.51-fold increased tendency for longevity, respectively (P=0.025 and 0.016). The haplotype results were strengthened by interaction analysis, which suggests an optimal model in which G190C and T2437C exert an interacting effect on longevity. No functional significance was observed between 190G and 190C alleles in both control and heat-inducible A549 cells (P>0.05). Taken together, our findings suggested that common genetic variants in Hsp70 family might contribute interactively to longevity the Xinjiang Hetian Uygur population. (Translational Research 2009;154:257-264)

Abbreviations: BMI - body mass index; Hsp70 - heat shock protein 70; PCR - polymerase chain reaction.

City (W.N.), and Grant XJ from the Science Fund of Shanghai Jiaotong University School of Medicine.

Submitted for publication March,; revision submitted August,; accepted for publication August,

Reprint requests: Wenquan Niu, Ruijin Hospital, Shanghai Institute of Hypertension, Ruijin Second Road, Shanghai, China; e-mail: niuwenquan@yahoo.cn, Zuheng Cheng, The First Hospital of Xinjiang Medical University; e-mail: zuheng-cheng@yahoo.com.cn or Changchun Qiu, Peking Union Medical College, Dong Dan San Tiao, Beijing, China; e-mail: qiucc@pumc.edu.cn, Tel: - /$ see front matter Mosby, Inc. All rights reserved. doi: /j.trsl.

We investigate the association of 3 variants (A1267G in HSPA1B, G190C in HSPA1A, and T2437C in HSPA1L) in heat shock protein 70 (Hsp70) with natural longevity in a Xinjiang Hetian Uygur population. Using a candidate gene approach and the promoter functional assessment, our findings suggested that common genetic variants in Hsp70 family might contribute interactively to longevity in the Xinjiang Hetian Uygur population.

We, therefore, focused on a population of Xinjiang Hetian Uygur ethnicity, whose percentage of centenarians is significantly greater than the average level of the whole country, to explore the association of 3 common genetic variants, including A1267G (rs1061581, exon 1), G190C (rs1043618, 5' UTR), and T2437C (rs2227956, exon 2) in 3 polymorphic genes (HSPA1B, HSPA1A, and HSPA1L) ofWe amplified a fragment of 627 base pairs (bp) and digested it into 354 bp and 273 bp in the presence of the HSPA1L 2437 T variant. After enzyme treatment, the reaction mixture was separated on 2% agarose gels. The primers and amplification conditions for the other two variants were according to a report by Wu et al.13. We confirmed the accuracy of our screening method by directly sequencing amplified DNA from randomly selected samples (10%), and found no difference between the two methods (data not shown).

To analyze the activity of HSPA1A, we cloned the promoter fragment (488 bp) encompassing the G190C variant into the pMD19-T Easy vector (TaKaRa) using forward primer: 5-CGG GGT ACC CGC CAT GGA GAC CAA CAC CC (contained KpnI restriction site) and reverse primer: 5- CCG CTC GAG CCG GTT CCC TGC TCT CTG TC. We confirmed construction by restriction analysis and DNA sequencing (BGI LifeTech Co., Ltd., Fremont, Calif). The fragment was then inserted upstream of the firefly luciferase reporter gene at the EcoRI site added between the KpnI/XhoI sites of the pGL3-basic vector (Promega Corporation, Madison, Wisc). We confirmed insert orientation in the HSPA1A-pGL3 construct by restriction assay.