Gene target specificity of the Super Elongation Complex (SEC) family: how HIV-1 Tat employs selected SEC members to activate viral transcription.
Bottom Line: Providing answers to these questions, our current study shows that AFF1 and AFF4 reside in separate SECs that display largely distinct gene target specificities.The functional difference between AFF1 and AFF4 in Tat-transactivation has been traced to a single amino acid variation between the two proteins, which causes them to enhance the affinity of Tat for P-TEFb, a key SEC component, with different efficiency.Finally, genome-wide analysis confirms that the genes regulated by AFF1-SEC and AFF4-SEC are largely non-overlapping and perform distinct functions.
Affiliation: Innovation Center of Cell Signaling Network, School of Pharmaceutical Sciences, Xiamen University, Xiamen 361005, Fujian, China Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.Show MeSH
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Mentions: To determine how the AFF1- and AFF4-SEC may function in a gene/activator-specific manner on a genome-wide scale, we expressed shAFF1 or shAFF4 in HeLa cells and performed RNA-seq to determine the downstream target genes regulated by the two types of SEC. The efficiency of KD as determined by qRT-PCR was high, with ∼80% of AFF1 and ∼90% of AFF4 depleted, respectively (Figure 6A). RNAs purified from the AFF1, AFF4 or the control GFP KD cells were prepared for single-end, high-throughput sequencing. The differentially expressed genes (DEGs) in response to AFF1 or AFF4 KD were identified by RankProd (P-value ≤ 0.05) (21). In total, 1517 and 1602 genes were differentially expressed in response to AFF1 and AFF4 KD, respectively (Figure 6B). Importantly, the majority of them (61.8% of the AFF1 KD- and 63.9% of the AFF4 KD-induced DEGs) were only responsive to AFF1 or AFF4 depletion, suggesting that to a large extent the AFF1- and AFF4-SEC control distinct subsets of target genes in vivo.
Affiliation: Innovation Center of Cell Signaling Network, School of Pharmaceutical Sciences, Xiamen University, Xiamen 361005, Fujian, China Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.