生物谷報(bào)道：p53的功能被多種翻譯后修飾機(jī)制所調(diào)控，包括Hdm2介導(dǎo)的泛素化作用，該作用可使其蛋白酶體降解。目前,，來自法國，德國,，西班牙等國家的學(xué)者發(fā)現(xiàn),，P53相關(guān)因子E4F1可以作為一個(gè)不典型的E3泛素鏈接酶，調(diào)控p53效應(yīng),，并且該過程不依賴于P53的降解,。這項(xiàng)新發(fā)現(xiàn)發(fā)表于最新一期的《Cell》雜志上。
E4F1是一種廣泛表達(dá)的鋅指蛋白,，最初被發(fā)現(xiàn)是病毒癌蛋白E1A的靶作用蛋白,。它可引起p53鉸鏈區(qū)的賴氨酸發(fā)生少量的泛素化，其作用機(jī)制不同于Hdm2,，后者是通過乙酰轉(zhuǎn)移酶PCAF乙?；囟ǖ陌械鞍?。因而，E4F1與PCAF對(duì)p53的調(diào)控作用是互斥的,。同時(shí),，依賴于E4F1的p53泛素化，伴隨著染色體的相關(guān),，以及p53信號(hào)通路的激活,，引起細(xì)胞周期停滯，而非凋亡,。
該研究證實(shí)了E4F1是p53的一個(gè)重要的翻譯后修飾調(diào)控因子,，并通過對(duì)p53功能的調(diào)控，影響著細(xì)胞的命運(yùn)：究竟是生長停滯還是凋亡,。這一發(fā)現(xiàn)對(duì)于未來進(jìn)一步研究p53的功能具有重要意義,。
Figure 1. E4F1 Exhibits Intrinsic Ubiquitin E3 Ligase Activity on p53 In Vitro and In Vivo
(A) Autoubiquitylation activity of E4F1. Autoradiograms of in vitro ubiquitylation assays performed under standard conditions in presence of in vitro translated (IVT) E4F1 labeled by 35S-methionine.
(B) Recombinant E4F1 stimulates p53 ubiquitylation in vitro. Autoradiograms of in vitro ubiquitylation assays performed with IVT 35S-labeled p53 and GST-E4F1 or GST.
(C) Autoradiograms of in vitro ubiquitylation assays performed under standard conditions with IVT 35S-labeled p53 and cellular E4F1 immunoprecipitated from E4F1- or mock-transfected U2OS cells.
(D) E4F1 stimulates K48-Ub branching. Autoradiograms of in vitro ubiquitylation assays performed with IVT 35S-labeled p53, baculovirus expressed E4F1 and Ub (WT), or Ub mutants bearing mutations of all (K0) or all but one of its lysine residues at the indicated position. Equal amounts of Ub mutants were used in each assay (Figure S1F).
(E) Cre-induced E4F1-GFP expression in U2OS cells stably expressing the LSL-E4F1 construct. (Upper panels) Western blot analyses of nuclear extracts prepared from LSL-E4F1 cells at the indicated time points after infection with Cre retrovirus, probed with anti-GFP, -E4F1, -p53 (DO1), -Cre, and -TBP (loading control) antibodies (Abs). (Lower panels) Analysis of E4F1-GFP expression by fluorescence microscopy 3 days after infection with Cre (+) or control (−) retroviruses. Cells are shown at 40× magnification.
(F) E4F1 stimulates ubiquitylation of endogenous p53 in vivo. Western blot analyses of Ub-conjugated proteins in LSL-E4F1 cells transiently transfected for 24 hr with a 6XHis-Ub expression vector 4 days after infection by Cre (+) or control (−) retroviruses. One percent of cellular extracts were probed for the presence of E4F1-GFP and total endogenous p53 using anti-GFP and anti-p53 (DO1) Abs (input). These cellular extracts, normalized to contain equal amounts of total p53, were loaded on nickel (Ni+)-NTA columns. Ni+-purified 6XHis-Ub-conjugated proteins were probed for the presence of Ub-p53 forms using an anti-p53 (DO1) Ab (Ni-purified). Neither cells nor cellular extracts were treated with proteasome inhibitors.
(G) Depletion of endogenous E4F1 impacts on endogenous p53 ubiquitylation in vivo. Western blot analyses of Ub-conjugated proteins in U2OS cells treated for 72 hr with control scramble or E4F1 SiRNAs. His-Ub-conjugated forms of endogenous p53 present in these cells were purified and analyzed as described in Figure 1F.
原文下載：
Cell November 17, 2006: 127 (4)
E4F1 Is an Atypical Ubiquitin Ligase that Modulates p53 Effector Functions Independently of Degradationp775
Laurent Le Cam, Laëtitia K. Linares, Conception Paul, Eric Julien, Matthieu Lacroix, Elodie Hatchi, Robinson Triboulet, Guillaume Bossis, Ayelet Shmueli, Manuel S. Rodriguez, Olivier Coux, and Claude Sardet
[Summary] [Full Text] [PDF] [Supplemental Data]
相關(guān)文獻(xiàn)：
A New Function for p53 Ubiquitination, Cell, Volume 127, Issue 4, 17 November 2006, Pages 675-677
Yuko Hirano1 and Ze'ev Ronai1,
SummaryPlus | Full Text + Links | PDF (252 K)
相關(guān)基因
Pubmed gene：
TP53
Official Symbol: TP53 and Name: tumor protein p53 (Li-Fraumeni syndrome) [Homo sapiens]
Other Aliases: LFS1, TRP53, p53
Other Designations: p53 tumor suppressor; tumor protein p53
Chromosome: 17; Location: 17p13.1
MIM: 191170
GeneID: 7157
E4F1
Official Symbol: E4F1 and Name: E4F transcription factor 1 [Homo sapiens]
Other Aliases: E4F, MGC99614
Other Designations: p120E4F
Chromosome: 16; Location: 16p13.3
MIM: 603022
GeneID: 1877
