与本result 相关的introduction对应为：

. Recently, we and others found that NLRP4/DTX4 and another E3 ligase TRAF-interacting protein (TRIP) induce the K48-linked  poly-ubiquitination of TBK1, leading to its proteasomal degradation (Cui et al., 2012; Zhang et al., 2012)

所以这里就是要说NLRP4信号体调节TBK1稳定性相关问题

开头说到core protein NLRP3,比较地引出了USP38，DTX4 AND TRIP三者与TBK1 的相互作用依赖于NLRP4。因此，根据发现，我们猜想：

NLRP4, together with USP38 and E3 ligases,might form the NLRP4 signalosome to control TBK1 stability and type I IFN signaling。

接下来我们分3个部分实验验证了猜想：

                                TBK1和NLRP4信号体成员间的相互作用

方法是：

infected MycTRIP- and GFP-DTX4-expressingcells with VSV, and then we immunoprecipitated different complexes from cells,collected at different time points post-infection with either TBK1 or NLRP4 antibodies.

通过免疫印迹分析结果发现：

NLRP4 and USP38 targeted to TBK1 at 6 hr post-infection, whereas DTX4 and TRIP interacted with TBK1 at a later time point (at 8 hr) (Figure 7A)

Importantly, USP38, but not DTX4 or TRIP, constitutively interacted with NLRP4 in unstimulated cells (Figure 7B).

Interaction between NLRP4 and USP38 was further increased after viral infection. Interactions between  NLRP4 and TRIP or DTX4 were detected at 6 or 8 hr, respectively, post-infection  (Figure 7B).

因此，所以结果都表明了：virus 入侵之后NLRP4的动态组成。

                         NLRP4信号转导体调节TBK1稳定性的分子机制

方法是：

generated NLRP4(-/-)andUSP38(-/-)HEK293T cells, respectively (Figures S6A and S6B).

我们发现：

USP38(-/-）cells and NLRP4(-/-）cells showed increasedISRE luciferase activity, ISG expression, and antiviral immunity after viralinfection (Figures S6C–S6F).

有了这个，我们进一步就确定NLRP4和USP38在TBK1和NLRP4信号转导体相互作用的角色：

USP38 deficiency does not affect the interaction of TBK1 with NLRP4, TRIP, or DTX4 (Figure 7C).By contrast, in NLRP4（-/-）cells, the interactions of TBK1 with USP38,TRIP, or DTX4 were completely abrogated (Figure 7D).

结果表明，在NLRP4信号转导体中，NLRP4是一个链接USP38,TRIP和DTX4这三者与TBK1的关键蛋白，而TBK1与NLRP4,TRIP和DTX4的相互作用却不依赖于USP38.

                            NLRP4信号转导体成员如何影响TBK1的ubiquitination

实验现象摆在这里：

observed increased levels of K33-linkedubiquitination and decreased levels of K48-linked ubiquitination of TBK1 in both USP38（-/-）and NLRP4（-/-）cells (Figures 7E and 7F). 

还有：

TRIP or DTX4 deficiency only affected K48-linked ubiquitination, but not K33-linked ubiquitination, of    TBK1 (Figures S7A and S7B).

结论告诉我们：

TBK1上K33关联的泛素化的消除需要NLRP4和USP38,同时K48相关的去泛素化需要E3连接酶DTX4和TRIP。

还有有趣的一个现象：

although NLRP4 or TRIP still interacted with TBK1 in USP38（-/-）cells (Figure 7C), they could not cause TBK1 degradation in USP38（-/-）cells even after VSV infection or IC  poly(I:C)treatment (Figures 7G,7H,S7C,and S7D). 

相对应的是：

Overexpression of WT USP38, but not the USP38 (CA/ HA) mutant,restored the function of NLRP4 to degrade TBK1 in USP38（-/-）cells (Figure 7I)。

这就是说，对于NLRP4信号转导体降解TBK1的全过程，USP38的去泛素化是灰常关键的。

而且，我们发现在NLRP4(-/-)细胞中USP38或者TRIP都不能降解TBK1.

因此，result 7的结论是：

NLRP4, USP38, TRIP, and DTX4 worked cooperatively as a signalosome to control TBK1 ubiquitination transition and degradation, thus inhibiting IFN-b signaling.。

So here comes a QUESTION:Whats the mechanism for these members  of the NLRP4 signalosome to control  TBK1 ubiquitination transition and degradation?