Novel reliable real-time PCR for differential detection of MSRVenv

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• 作者：Giuseppe Mameli ; Luciana Poddighe ; Vito Astone ; Giuseppe Delogu ; Giannina Arru ; Stefano Sotgiu ; Caterina Serra ; Antonina Dolei
• 关键词：Human endogenous retroviruses ; Multiple sclerosis ; Multiple sclerosis-associated retrovirus ; Syncytin-1 ; HERV-W ; Discriminatory real-time PCR
• 刊名：Journal of Virological Methods
• 出版年：2009
• 出版时间：October 2009
• 年：2009
• 卷：161
• 期：1
• 页码：98-106
• 全文大小：636 K

文摘

Two components of the HERV-W family of human endogenous retroviruses are activated during multiple sclerosis (MS) and proposed immunopathogenic co-factors: MSRV (MS-associated retrovirus), and ERVWE1 (whose env protein, syncytin-1, reaches the plasma membrane). MSRVenv and syncytin-1 are closely related, and difficult to distinguish each other. The sequences of extracellular MSRVenv and of syncytin-1 available in GenBank were compared with those found in MS patients and controls of the cohort under study. With respect to syncytin-1, MSRVenv sequences have a 12-nucleotide insertion in the trans-membrane moiety. Based on this insertion, discriminatory real-time PCR assays were developed, that can amplify selectively either MSRVenv or syncytin-1. The data of MS patients and controls indicated that MSRV and ERVWE1 are both expressed in the brain of MS patients, while only MSRV is present in the blood; MSRV was released in culture by PBMCs of MSRV-producer individuals. These cells expressed the complete MSRVenv gene in the absence of syncytin-1 expression, up to the final, fully glycosylated envelope protein product, since western blot staining with anti-HERV-Wenv antibody detected two bands of the same molecular weight (73 and 61 kDa) of the fully glycosylated and partially glycosylated HERV-Wenv uncleaved proteins. Beyond MSRVenv DNA copy numbers were more abundant in MS patients than in healthy humans, while syncytin-1 were unchanged. These findings reinforce the link between MSRV and MS.