Mendoza Heidi, Campbell David G, Burness Kerry, Hastie James, Ronkina Natalia, Shim Jae-Hyuck, Arthur J Simon C, Davis Roger J, Gaestel Matthias, Johnson Gary L, Ghosh Sankar, Cohen Philip
MRC Protein Phosphorylation Unit, University of Dundee, Dundee DD1 5EH, Scotland, UK.
Biochem J. 2008 Feb 1;409(3):711-22. doi: 10.1042/BJ20071149.
The protein kinase TAK1 (transforming growth factor-beta-activated kinase 1), which has been implicated in the activation of MAPK (mitogen-activated protein kinase) cascades and the production of inflammatory mediators by LPS (lipopolysaccharide), IL-1 (interleukin 1) and TNF (tumour necrosis factor), comprises the catalytic subunit complexed to the regulatory subunits, termed TAB (TAK1-binding subunit) 1 and either TAB2 or TAB3. We have previously identified a feedback-control mechanism by which p38alpha MAPK down-regulates TAK1 and showed that p38alpha MAPK phosphorylates TAB1 at Ser(423) and Thr(431). In the present study, we identified two IL-1-stimulated phosphorylation sites on TAB2 (Ser(372) and Ser(524)) and three on TAB3 (Ser(60), Thr(404) and Ser(506)) in human IL-1R cells [HEK-293 (human embryonic kidney) cells that stably express the IL-1 receptor] and MEFs (mouse embryonic fibroblasts). Ser(372) and Ser(524) of TAB2 are not phosphorylated by pathways dependent on p38alpha/beta MAPKs, ERK1/2 (extracellular-signal-regulated kinase 1/2) and JNK1/2 (c-Jun N-terminal kinase 1/2). In contrast, Ser(60) and Thr(404) of TAB3 appear to be phosphorylated directly by p38alpha MAPK, whereas Ser(506) is phosphorylated by MAPKAP-K2/MAPKAP-K3 (MAPK-activated protein kinase 2 and 3), which are protein kinases activated by p38alpha MAPK. Studies using TAB1(-/-) MEFs indicate important roles for TAB1 in recruiting p38alpha MAPK to the TAK1 complex for the phosphorylation of TAB3 at Ser(60) and Thr(404) and in inhibiting the dephosphorylation of TAB3 at Ser(506). TAB1 is also required to induce TAK1 catalytic activity, since neither IL-1 nor TNFalpha was able to stimulate detectable TAK1 activity in TAB1(-/-) MEFs. Surprisingly, the IL-1 and TNFalpha-stimulated activation of MAPK cascades and IkappaB (inhibitor of nuclear factor kappaB) kinases were similar in TAB1(-/-), MEKK3(-/-) [MAPK/ERK (extracellular-signal-regulated kinase) kinase kinase 3] and wild-type MEFs, suggesting that another MAP3K (MAPK kinase kinase) may mediate the IL-1/TNFalpha-induced activation of these signalling pathways in TAB1(-/-) and MEKK3(-/-) MEFs.
蛋白激酶TAK1（转化生长因子-β激活激酶1）与丝裂原活化蛋白激酶（MAPK）级联反应的激活以及脂多糖（LPS）、白细胞介素1（IL-1）和肿瘤坏死因子（TNF）诱导的炎症介质产生有关，它由与调节亚基（称为TAK1结合亚基（TAB）1以及TAB2或TAB3）复合的催化亚基组成。我们之前鉴定出一种反馈控制机制，通过该机制p38α MAPK下调TAK1，并表明p38α MAPK使TAB1的丝氨酸423和苏氨酸431位点磷酸化。在本研究中，我们在人IL-1R细胞[稳定表达IL-1受体的人胚肾（HEK-293）细胞]和小鼠胚胎成纤维细胞（MEF）中鉴定出TAB2上两个IL-1刺激的磷酸化位点（丝氨酸372和丝氨酸524）以及TAB3上三个位点（丝氨酸60、苏氨酸404和丝氨酸506）。TAB2的丝氨酸372和丝氨酸524不会被依赖p38α/β MAPK、细胞外信号调节激酶1/2（ERK1/2）和c-Jun氨基末端激酶1/2（JNK1/2）的信号通路磷酸化。相反，TAB3的丝氨酸60和苏氨酸404似乎直接被p38α MAPK磷酸化，而丝氨酸506则被p38α MAPK激活的蛋白激酶MAPKAP-K2/MAPKAP-K3磷酸化。使用TAB1基因敲除的MEF进行的研究表明，TAB1在将p38α MAPK募集到TAK1复合物以实现TAB3丝氨酸60和苏氨酸404位点的磷酸化以及抑制TAB3丝氨酸506位点的去磷酸化过程中发挥重要作用。诱导TAK1催化活性也需要TAB1，因为在TAB1基因敲除的MEF中，IL-1和TNFα均无法刺激检测到的TAK1活性。令人惊讶的是，在TAB1基因敲除的、MEKK3基因敲除的（MAPK/ERK激酶激酶3）和野生型MEF中，IL-1和TNFα刺激的MAPK级联反应和核因子κB抑制蛋白激酶（IkappaB）的激活情况相似，这表明可能有另一种MAPK激酶激酶（MAP3K）介导了IL-1/TNFα在TAB1基因敲除的和MEKK3基因敲除的MEF中诱导的这些信号通路的激活。
