The Super Elongation Complex (SEC), containing transcription elongation activators/coactivators P-TEFb, ELL2, AFF4/1, ENL, and AF9, is recruited by HIV-1 Tat and mixed lineage leukemia (MLL) proteins to activate the expression of HIV-1 and MLL-target genes, respectively. In the absence of Tat and MLL, however, it is unclear how SEC is targeted to RNA polymerase (Pol) II to stimulate elongation in general. Furthermore, although ENL and AF9 can bind the H3K79 methyltransferase Dot1L, it is unclear whether these bindings are required for SEC-mediated transcription. Here, we show that the homologous ENL and AF9 exist in separate SECs with similar but nonidentical functions. ENL/AF9 contacts the scaffolding protein AFF4 that uses separate domains to recruit different subunits into SEC. ENL/AF9 also exists outside SEC when bound to Dot1L, which is found to inhibit SEC function. The YEATS domain of ENL/AF9 targets SEC to Pol II on chromatin through contacting the human Polymerase-Associated Factor complex (PAFc) complex. This finding explains the YEATS domain's dispensability for leukemogenesis when ENL/AF9 is translocated to MLL, whose interactions with PAFc and DNA likely substitute for the PAFc/chromatin-targeting function of the YEATS domain.A ccumulating evidence has implicated the elongation stage of RNA polymerase II (Pol II) transcription as a major ratelimiting step for the expression of a large number of metazoan genes, especially those that control cell growth, renewal, and differentiation (1-3). During elongation, the processivity of Pol II is regulated by a set of transcription factors, which had been thought to exist as separate entities and impact on the Pol II elongation complex independently of one another. However, recent data from us and others indicate that at least two well defined transcription elongation factors of different classes reside in a single multisubunit complex termed SEC [super elongation complex, (4)] to cooperatively activate transcription (4-6).The first elongation factor found in SEC is human positive transcription elongation factor b (P-TEFb). Consisting of CDK9 (Cyclin-dependent Kinase 9) and cyclin T1 (CycT1), P-TEFb functions by phosphorylating the C-terminal domain (CTD) of the largest subunit of Pol II and negative elongation factors DSIF and NELF. These events antagonize the actions of the negative factors, release Pol II from promoter-proximal pausing, and trigger the production of full-length mRNA transcripts (7,8). The second elongation factor in SEC is ELL2, which promotes elongation by keeping the 3′ OH of nascent mRNA in alignment with the catalytic site to prevent Pol II backtracking (9). Besides P-TEFb and ELL2, SEC also contains transcription factors/coactivators ENL, AF9, AFF4 (AF5q31), AFF1 (AF4), and probably others (4-6). Among these, AFF4 is known to mediate the interaction between ELL2 and P-TEFb and maintain the integrity of SEC (5). While AFF1 has been shown to interact with AFF4 (10), it remains to be seen whether the interaction between these two homologo...

Super Elongation Complexes (SECs) contain two different transcription elongation factors, P-TEFb and ELL1/2, linked by the scaffolding protein AFF4 or AFF1. They stimulate the expression of both normal and disease-related genes, especially those of HIV or involved in leukemogenesis. Among all SECs subunits, ELL2 is stoichiometrically limiting and uniquely regulated at the level of protein stability. Here we identify the RING domain protein Siah1, but not the homologous Siah2, as the E3 ubiquitin ligase for ELL2 polyubiquitination and proteasomal degradation. Siah1 cannot access and ubiquitinate ELL2 bound to AFF4, although at high concentrations, it also degrades AFF4/1 to destroy SECs. Prostratin and HMBA, two well-studied activators of HIV transcription and latency, enhance ELL2 accumulation and SECs formation largely through decreasing Siah1 expression and ELL2 polyubiquitination. Given its importance in formation of SECs, the Siah1 ubiquitination pathway provides a fresh avenue for developing strategies to control disease-related transcription.

SUMMARY Efficient chemotaxis requires rapid coordination between different parts of the cell in response to changing directional cues. Here we investigate the mechanism of front-rear coordination in chemotactic neutrophils. We find that changes in the protrusion rate at the cell front are instantaneously coupled to changes in retraction at the cell rear, while myosin II accumulation at the rear exhibits a reproducible 9-15 sec lag. In turning cells, myosin II exhibits dynamic side-to-side relocalization at the cell rear in response to turning of the leading edge, and facilitates efficient turning by rapidly re-orienting the rear. These manifestations of front-rear coupling can be explained by a simple quantitative model incorporating reversible actin-myosin interactions with a rearward-flowing actin network. Finally, the system can be tuned by the degree of myosin regulatory light chain (MRLC) phosphorylation, which appears to be set in an optimal range to balance persistence of movement and turning ability.

The cytoplasm of a living cell is a dynamic environment through which intracellular components must move and mix. In motile, rapidly deforming cells such as human neutrophils, bulk cytoplasmic flow couples cell deformation to the transport and dispersion of cytoplasmic particles. Using particle-tracking measurements in live neutrophil-like cells, we demonstrate that fluid flow associated with the cell deformation contributes to the motion of small acidic organelles, dominating over diffusion on timescales above a few seconds. We then use a general physical model of particle dispersion in a deforming fluid domain to show that transport of organelle-sized particles between the cell periphery and the bulk can be enhanced by dynamic deformation comparable to that observed in neutrophils. Our results implicate an important mechanism contributing to organelle transport in these motile cells: cytoplasmic flow driven by cell shape deformation.