Muckenfuss Heide, Hamdorf Matthias, Held Ulrike, Perković Mario, Löwer Johannes, Cichutek Klaus, Flory Egbert, Schumann Gerald G, Münk Carsten
Division of Medical Biotechnology, Paul-Ehrlich-Institut, Paul-Ehrlich-Strasse 51-59, D-63225 Langen, Germany.
Section Pr2/Retroelements, Paul-Ehrlich-Institut, Paul-Ehrlich-Strasse 51-59, D-63225 Langen, Germany.
J Biol Chem. 2006 Aug 4;281(31):22161-22172. doi: 10.1074/jbc.M601716200. Epub 2006 May 30.
The human cytidine deaminase family APOBEC3 represents a novel group of proteins in the field of innate defense mechanisms that has been shown to be active against a variety of retroviruses. Here we examined whether members of the APO-BEC3 family have an impact on retrotransposition of human long interspersed nuclear elements (LINE-1s or L1s). Using a retrotransposition reporter assay in HeLa cells, we demonstrate that in the presence of transiently transfected APOBEC3A, L1 retrotransposition frequency was reduced by up to 85%. Although APOBEC3G and -3H did not influence L1 retrotransposition notably, expression of APOBEC3B, -3C, and -3F inhibited transposition by approximately 75%. Although reverse transcription of L1s occurs in the nucleus and APOBEC3 proteins are believed to act via DNA deamination during reverse transcription, activity against L1 retrotransposition was not correlated with nuclear localization of APOBEC3s. We demonstrate that APOBEC3C and APOBEC3B were endogenously expressed in HeLa cells. Accordingly, down-regulation of APOBEC3C by RNA interference enhanced L1 retrotransposition by approximately 78%. Sequence analyses of de novo L1 retrotransposition events that occurred in the presence of overexpressed APOBEC3 proteins as well as the analyses of pre-existing endogenous L1 elements did not reveal an enhanced rate of G-to-A transitions, pointing to a mechanism independent of DNA deamination. This study presents evidence for a role of host-encoded APOBEC3 proteins in the regulation of L1 retrotransposition.
人类胞苷脱氨酶APOBEC3家族代表了天然防御机制领域中的一组新型蛋白质，已证明其对多种逆转录病毒具有活性。在此，我们研究了APOBEC3家族成员是否对人类长散在核元件（LINE-1或L1）的逆转座有影响。通过在HeLa细胞中进行逆转座报告基因检测，我们证明在瞬时转染APOBEC3A的情况下，L1逆转座频率降低了85%。虽然APOBEC3G和-3H对L1逆转座没有显著影响，但APOBEC3B、-3C和-3F的表达使转座受到约75%的抑制。虽然L1的逆转录发生在细胞核中，并且APOBEC3蛋白被认为在逆转录过程中通过DNA脱氨发挥作用，但对L1逆转座的活性与APOBEC3的核定位无关。我们证明APOBEC3C和APOBEC3B在HeLa细胞中内源性表达。因此，通过RNA干扰下调APOBEC3C可使L1逆转座增加约78%。对在过表达APOBEC3蛋白情况下发生的新生L1逆转座事件的序列分析以及对预先存在的内源性L1元件的分析均未发现G到A转换率增加，这表明存在一种独立于DNA脱氨的机制。这项研究为宿主编码的APOBEC3蛋白在L1逆转座调控中的作用提供了证据。
