Exploring Genetic Factors Involved in Huntington Disease Age of Onset: E2F2 as a New Potential Modifier Gene

Abstract

Age of onset (AO) of Huntington disease (HD) is mainly determined by the length of the CAG repeat expansion (CAGexp) in exon 1 of the HTT gene. Additional genetic variation has been suggested to contribute to AO, although the mechanism by which it could affect AO is presently unknown. The aim of this study is to explore the contribution of candidate genetic factors to HD AO in order to gain insight into the pathogenic mechanisms underlying this disorder. For that purpose, two AO definitions were used: the earliest age with unequivocal signs of HD (earliest AO or eAO), and the first motor symptoms age (motor AO or mAO). Multiple linear regression analyses were performed between genetic variation within 20 candidate genes and eAO or mAO, using DNA and clinical information of 253 HD patients from REGISTRY project. Gene expression analyses were carried out by RT-qPCR with an independent sample of 35 HD patients from Basque Country Hospitals. We found suggestive association signals between HD eAO and/or mAO and genetic variation within the E2F2, ATF7IP, GRIN2A, GRIN2B, LINC01559, HIP1 and GRIK2 genes. Among them, the most significant was the association between eAO and rs2742976, mapping to the promoter region of E2F2 transcription factor. Furthermore, rs2742976 T allele patient carriers exhibited significantly lower lymphocyte E2F2 gene expression, suggesting a possible implication of E2F2-dependent transcriptional activity in HD pathogenesis. Thus, E2F2 emerges as a new potential HD AO modifier factor.

Fig 1. RT-qPCR analysis of E2F2 gene expression in HD patients, according to E2F2 rs2742976 genotype.

Two methods were used. In Taqman assay, the expression of E2F2 gene was analyzed in 31 samples (N _TT = 4; N _GT = 12, N _GG = 15) with Hs00918089_m1 Taqman probe; the expression values were normalized respect to expression of B2M and YWHAZ reference genes. In SYBR Green assay, the E2F2 gene expression was estimated in 31 samples (N_TT = 5; N_GT = 14, N_GG = 12); the expression values were normalized to expression of UBC and YWHAZ reference genes. Results are expressed as fold over respective GG individuals. Asterisk denotes statistically significant differences (P<0.05) between GG and any other group, according to DataAssist software analysis (T-test) or REST software analysis (Pair Wise Reallocation Randomization test).