InteractiveFly: GeneBrief

Su(Tpl): Biological Overview | References

BIOLOGICAL OVERVIEW

Many developmentally regulated genes contain a poised RNA polymerase II (Pol II) at their promoters under conditions where full-length transcripts are undetectable. It has been proposed that the transcriptional activity of such promoters is regulated at the elongation stage of Pol II transcription. In Drosophila, the heat-shock loci expressing the Hsp70 genes have been used as a model for the regulation of the transcriptional activity of poised Pol II. Drosophila ELL [dELL; Su(Tpl)] is a Pol II elongation factor capable of stimulating the rate of transcription both in vivo and in vitro. Although ELL and the elongation factor Elongin A have indistinguishable effects on RNA polymerase in vitro, the loss-of-function studies indicate that these proteins are not redundant in vivo. This study used RNAi to investigate the physiological properties of dELL and a dELL-associated factor (dEaf) in a living organism. Both ELL and Eaf are essential for fly development. dELL is recruited to heat shock loci upon induction, and its presence with Pol II at such loci is required for proper heat-shock gene expression. Consistent with a role in elongation, dELL knockdown reduces the levels of phosphorylated Pol II at heat-shock loci. This study implicates dELL in the expression of loci regulated by Pol II elongation (Smith, 2008).

Efficient transcription by RNA polymerase II (Pol II) is an intricate process that requires multiple contacts with the DNA template and nascent RNA that inevitably leads to frequent stalling during the transcription of a gene. The average rate of transcription by Pol II in vivo is an order of magnitude higher than that obtained in vitro despite additional impediments, such as traversing through nucleosomes. Using biochemical approaches, two Pol II elongation factors, Eleven nineteen lysine-rich leukemia (ELL) and Elongin A, were isolated from cell extracts as factors capable of stimulating Pol II activity by suppressing transient pausing. Despite similar in vitro activities, the Drosophila orthologs of ELL and Elongin A are each essential for development (Eissenberg, 2002; Gerber, 2004). This observation indicates that their in vivo activity is not redundant (Smith, 2008).

Recent genome-wide studies have found a large number of developmentally regulated genes that contain a paused Pol II at their promoters (Muse, 2007; Zeitlinger, 2007). Therefore, it has been proposed that the transcriptional activity of such poised Pol IIs is regulated at the level of transcription elongation. The classic model for studying genes regulated by promoter-proximal paused polymerase is Hsp70 gene induction in Drosophila. Previous studies have shown that several Pol II elongation factors are rapidly recruited to the Hsp70 genes after heat shock. Although much work has been done on the role of these factors in gene regulation in cultured cells, less is known about the role of these factors in the regulation of heat-shock gene expression in the whole organism. Although there are several mutants in the gene encoding Drosophila ELL (dELL), all of these alleles are embryonic lethal (Eissenberg, 2002). Therefore, it was possible to use these alleles to further characterize the role of the elongation factor ELL in the regulation of the transcriptional activity of poised Pol II and Hsp70 loci. To test the role for dELL in gene expression, RNAi was used to reduce expression levels of both dELL and dELL-associated factor (dEaf) expression levels during development, and the in vivo effect of their reduction on transcription and development was examined. It was found that knockdown of dELL and dEaf results in lethality. Furthermore, knockdown of these elongation factors results in reduced Hsp70 transcript accumulation after heat shock. Immunolocalization of phosphorylated Pol II in heat-shocked dELL knockdown salivary glands demonstrates reduced levels of the elongating form of Pol II at the Hsp70 loci in the absence of dELL. These studies demonstrate that dELL is essential for full induction of heat-shock gene expression and are consistent with a role for dELL in Pol II elongation. These findings provide a role for an RNA Pol II elongation factor in the transcriptional regulation of poised Pol II (Smith, 2008).

dELL has been shown to be essential; homozygous mutant clones do not survive in the eye and homozygotes for loss-of-function alleles die at the end of embryogenesis or in early first instar. To investigate the role of dELL in transcription in flies, dELL was knocked down by RNAi, which typically reduces, but does not eliminate, the targeted gene products. A 600-bp portion of the dELL coding region was inserted into a P-element vector that drives the expression of dsRNA through two convergent Gal4 UAS promoters that flank the insert. Several transgenic lines were generated and tested for effects on viability by crossing to an Actin5C-Gal4 driver line that expresses yeast Gal4 under the cytoplasmic actin promoter. All eight dELL RNAi lines show significant loss of viability when expressed under this driver. When adult escapers were obtained, very few males were observed, indicating that males are more susceptible to loss of dELL. Greater numbers of females than males were observed at the third instar larval stage, indicating that males are dying earlier than females. A significant genome-wide reduction of dELL protein is observed by immunofluorescence analysis of dELL RNAi larval polytene chromosomes (Smith, 2008).

Through two-hybrid analysis, two interacting partners of ELL have been characterized in humans, Eaf1 and Eaf2 (Simone, 2001; Simone, 2003). Eaf1 and Eaf2 are highly related and can stimulate the elongation activity of ELL in vitro (Kong, 2005). Recently, the association of Eaf with ELL was shown to be evolutionarily conserved, with the finding that Schizosaccharomyces pombe homologs SpEaf and SpELL directly interact with each other (Banks, 2007). Additionally, SpEaf enhances the stimulation by SpELL of Pol II transcription in vitro (Banks, 2007). Because Drosophila also has a single Eaf homolog, RNAi was used to knock down dEaf levels and assessed the viability of dEaf-knockdown flies in six different transgenic RNAi lines. In all lines, significant reductions were observed in the number of adult progeny of RNAi-expressing flies compared with control siblings. In addition, a consistent reduction in the male-female sex ratio was observed for dEaf RNAi, suggesting that the male-enhanced lethal phenotype (not observed for other elongation factors) is due to loss of a dELL-dEaf complex (Smith, 2008).

To test for the effectiveness of the RNAi knockdowns, dELL and dEaf mRNA levels were measured in knockdown larvae and their control siblings. Significant reductions in dELL transcripts are observed in the dELL RNAi larvae. dELL transcripts, as measured by RT-PCR, are not reduced by RNAi to the same level as dELL protein, as assessed by immunofluorescence on polytene chromosomes. Previously, it was observed that knockdown of dRTF1 by RNAi was more effective at the protein than the RNA levels presumably because the long dsRNAs produced are processed as miRNAs and interfere with translation. Because dELL is nested in an intron of the gene encoding the chromatin remodeling enzyme dMi-2, transcript levels for this gene were measured and no reduction was found of dMi-2 RNA in dELL RNAi larvae. Additionally, it was found that dEaf RNA levels are reduced in dEaf RNAi larvae. Interestingly, a significant increase in dELL levels is observed in dEaf RNAi larvae, possibly compensating for the lower dEaf levels (Smith, 2008).

dELL was previously shown to be recruited to heat-shock genes upon heat shock (Gerber, 2001). To determine whether dELL is required for heat-shock gene expression, the levels of Hsp70 transcripts after heat shock were compared in dELL knockdown larvae and their control siblings. By immunofluorescence analysis, little or no dELL is seen at the Hsp70 gene after heat shock in dELL knockdown larvae, whereas the control siblings without the Gal4 driver showed the expected recruitment of dELL to the Hsp70 gene. Northern blot analysis showed reduced levels of Hsp70 mRNA levels in the dELL RNAi larvae. A similar analysis was done with dEaf RNAi larvae, and reduced Hsp70 mRNA also occurs after heat shock, although the deficit was less than observed for the dELL RNAi larvae. Similar results were observed when Hsp70 levels were measured by RT-PCR, showing greater reductions in Hsp70 RNA levels in dELL RNAi than dEaf RNAi larvae (Smith, 2008).

Chromosomal levels of dELL are markedly reduced in the absence of Cdk9, the catalytic subunit of the Pol II C-terminal domain (CTD) kinase PTEF-B (Eissenberg, 2007). To determine whether dELL knockdown affects the recruitment of Pol II to the Hsp70 genes, dELL knockdown and control polytene chromosomes were probed with antibodies to the Ser-2-phosphorylated, elongating form of Pol II. Lower levels of Ser-2-phosphorylated Pol II were consistently observed at the Hsp70 heat-shock loci in dELL-knockdown larvae, suggesting a close link between dELL function and phosphorylation of the Pol II CTD (Smith, 2008).

ELL belongs to a class of transcription elongation factors that have been shown to stimulate the K_m and/or V_max of RNA Pol II in vitro by alleviating pausing on a purified DNA template. Another member of this class is Elongin A and its Drosophila ortholog dEloA (Shilatifard, 2003; Eissenberg, 2002; Gerber, 2004). From the present and previous studies, it is clear that both dELL and dEloA localize to the Hsp70 gene upon heat shock, and each is required for full levels of heat-shock gene expression, suggesting that the in vivo roles of these elongation factors in Hsp70 gene transcription are not redundant (Gerber, 2005). Similarly, it was observed that the knockdown phenotypes of these two proteins can be unique, such as the enhanced male lethality in dELL RNAi larvae. How could both elongation factors be redundant in vitro, yet nonredundant in vivo? The in vitro studies were performed on naked DNA templates, whereas the chromatin environment of RNA Pol II-transcribed genes can provide additional challenges to the polymerase. Each of these elongation factors has its own interaction partners and may be recruited to distinct states of the polymerase, such as initiating, elongating, or stalled polymerase. Consistent with this view, knockdown of dELL, but not dEloA, results in decreased levels of Ser-2-phosphorylated Pol II at the Hsp70 and other loci. Interestingly, the chromosomal targeting of dELL, but not dEloA, is dramatically reduced by the knockdown of CDK9, the Pol II CTD kinase, suggesting that dELL and dEloA are recruited to genes by distinct mechanisms. Fine mapping of dELL and dEloA on the well characterized Hsp70 gene at different time points after activation could clarify the distinct roles for these enzymes (Smith, 2008).

The lesser effect of dEaf knockdown on Hsp70 gene induction could be indicative of a requirement of dEaf for optimal function of dELL, whereas dELL can partially function without dEaf. Indeed, in vitro transcription studies have demonstrated that human Eaf proteins, in combination with ELL, stimulate transcription elongation by Pol II above the levels obtained with ELL alone (Kong, 2005). In dEaf RNAi larvae, it was observed that dELL levels are increased, conceivably as a cellular response to increased pausing resulting from lower dEaf levels (Smith, 2008).

Previous work on the function of dELL made use of alleles of the Su(Tpl) locus, which encodes dELL (Eissenberg, 2002). All known Su(Tpl) alleles are embryonic lethal. In contrast, RNAi of dELL allows survival to the larval or adult stages depending on the insertion line of the dsRNA construct. Interestingly, the few 'escaper' dELL RNAi adults are overwhelmingly female. As seen with the heat-shock defect, the difference in male and female viability is less in dEaf RNAi flies than in dELL RNAi flies, consistent with dEaf enhancing, but not being absolutely required for, dELL function. A previous study showed that males express much higher levels of a dELL transcript than females, although the functional significance of this difference has not been investigated (Gerber, 2001). One hypothesis is that dELL is needed in males as part of the process of X chromosome dosage compensation; Drosophila dosage compensation factors are thought to enhance transcription elongation of X-linked genes in males, and loss of any of these factors leads to male-specific lethality. In addition, reduced levels of several global chromatin regulators, including the supercoiling factor, Jil-1 H3 kinase, heterochromatin protein HP1, and the chromatin remodeler ISWI, have been reported to differentially affect the survival of males and/or the morphology of the X chromosome. However, in dELL knockdowns, MSL localization and the male polytene X chromosome morphology appears similar in dELL knockdown male larvae and their control brothers. Whether there are specific defects in dosage compensation of X-linked genes may be an interesting avenue for future investigations. Alternative explanations for a male-enhanced lethality also should be considered. For example, Drosophila males differ from females not just in having one less X chromosome, but also in carrying a Y chromosome, which comprises ~12% of the male genome. A number of genes are male-lethal due to the presence of the mostly heterochromatic Y chromosome, including modulators of position effect variegation, such as the Su(var)3-3 gene that encodes the histone demethylase LSD1, the uncharacterized Su(var)2-1, as well as the HP1-interacting protein Bonus (dTIF1), an enhancer and suppressor of position-effect variegation. For Su(var)2-1 and Bonus, the Y-lethal effect is not Y-specific but can be phenocopied by other sources of heterochromatin. A role for dELL in the regulation of heterochromatin is unknown but could conceivably be required for the expression of heterochromatin components (Smith, 2008).

Cdk9 is an essential kinase in Drosophila that is required for heat shock gene expression, histone methylation and elongation factor recruitment

Phosphorylation of the large RNA Polymerase II subunit C-terminal domain (CTD) is believed to be important in promoter clearance and for recruiting protein factors that function in messenger RNA synthesis and processing. P-TEFb is a protein kinase that targets the (CTD). The goal of this study was to identify chromatin modifications and associations that require P-TEFb activity in vivo. The catalytic subunit of P-TEFb, Cdk9, was knocked down in Drosophila using RNA interference. Cdk9 knockdown flies die during metamorphosis. Phosphorylation at serine 2 and serine 5 of the CTD heptad repeat were both dramatically reduced in knockdown larvae. Hsp 70 mRNA induction by heat shock was attenuated in Cdk9 knockdown larvae. Both mono- and trimethylation of histone H3 at lysine 4 were dramatically reduced, suggesting a link between CTD phosphorylation and histone methylation in transcribed chromatin in vivo. Levels of the chromo helicase protein CHD1 were reduced in Cdk9 knockdown chromosomes, suggesting that CHD1 is targeted to chromosomes through P-TEFb-dependent histone methylation. Dimethylation of histone H3 at lysine 36 was significantly reduced in knockdown larvae, implicating CTD phosphorylation in the regulation of this chromatin modification. Binding of the RNA Polymerase II elongation factor ELL was reduced in knockdown chromosomes, suggesting that ELL is recruited to active polymerase via CTD phosphorylation (Eissenberg, 2007).

Cdk9, the catalytic subunit of P-TEFb, is highly conserved among eukaryotes. The yeast kinases Ctk1 and Bur1 are both homologs of Cdk9, and both are CTD kinases in Drosophila, although loss of Bur1 has no effect on CTD phosphorylation yeast. Bur1 is essential but Ctk1 is not (Eissenberg, 2007).

RNAi knockdown of Cdk9 in transgenic flies results in lethality at the pupal stage. This is considerably later than the embryonic lethality reported for C. elegans RNAi knockdown of Cdk9. While this difference could reflect differences in the requirements for Cdk9 in these organisms, it is more likely that differences in timing or efficiency of RNAi, Cdk9 protein turnover and/or maternal Cdk9 loading accounts for the much later lethality in knockdown flies. Nevertheless, these results confirm and extend the finding that P-TEFb is essential in metazoan development (Eissenberg, 2007).

In contrast, Cdk9 homologs in fission yeast and Neurospora are not essential. Since CTD phosphorylation has been linked to promoter clearance, pre-mRNA processing and chromatin modification, it is not possible to say what aspect of P-TEFb activity is essential in metazoa. RNAi knockdown of the Drosophila Cdk9 in cultured cells causes arrest of the cell cycle at the G1-S transition, implicating this kinase in cell cycle control. It is unlikely that cell cycle arrest is causing the lethality in knockdown flies, since cell cycle mutations in Drosophila generally are associated with reduced or missing imaginal discs, and the discs in Cdk9 knockdown larvae appear overtly normal. The finding that Hsp70 transcripts are reduced in Cdk9 knockdown larvae is consistent with the reduced Hsp70 transcription previously reported in Cdk9 RNAi cultured cells. Hsp 70 is not essential in Drosophila, but the effects on Hsp70 suggest that defects in gene expression could underlie the essential requirement for Cdk9 in Drosophila development (Eissenberg, 2007).

Cdk9 knock-down flies show dramatic reductions in both serine 2 and serine 5 phosphorylation. In contrast, flavopiridol treatment of cultured cells has been found to selectively reduce serine 2 phosphorylation. The significance of this difference is unclear, but could reflect differences in experimental protocol. For example, flavopiridol treatments were limited to 15-20 min, while RNAi knockdown third instar larvae are subject to knockdown conditions for several days before assay. Longer periods of Cdk9 inactivation may be required for reduction in serine 5 phosphorylation. Alternatively, it is possible that knockdown of Cdk9 protein levels results in inhibition of TFIIH, the other known CTD kinase. Regardless of the mechanism, the RNAi knockdown clearly results in reduced phosphorylation of the CTD, enabling a test of the consequences of loss of CTD phosphorylation on chromatin modification and recruitment of RNA Polymerase II-associated factors (Eissenberg, 2007).

Loss of CTD phosphorylation in Cdk9 knockdown larvae is associated with reduced binding of the RNA Polymerase II elongation factor ELL genome-wide. ELL is broadly co-localized with phosphorylated RNA Polymerase II on polytene chromosomes, and is rapidly recruited to heat shock loci after a brief heat shock. These results suggest that the efficient recruitment of ELL to transcribed loci requires CTD phosphorylation. Whether this reflects a direct interaction of ELL with the CTD is unknown (Eissenberg, 2007).

Despite the fact that Elongin A affects the same kinetic parameter in RNA Polymerase II catalysis as ELL, Elongin A binding is not reduced by loss of CTD phosphorylation. As with ELL, the nature of Elongin A binding to RNA Polymerase II is unknown, but these observations suggest their binding can be distinguished by sensitivity to the phosphorylation state of the CTD. Since no increase of Elongin A was observed under conditions of reduced ELL binding, it seems unlikely that ELL and Elongin A compete for RNA Polymerase II binding (Eissenberg, 2007).

Spt4 and Spt5 are subunits of DSIF, which is implicated in the regulation of RNA Polymerase II elongation. Previous work suggested that reduced serine 2 phosphorylation of the RNA Polymerase II CTD has no effect on Spt5 recruitment to a heat shock gene in cultured cells (Ni, 2004). In Cdk9 knockdown flies, in which both serine 2 and 5 phosphorylation are reduced, the chromosomal distribution of Spt5 is unchanged genome-wide. This is consistent with previous reports that Spt5 interacts with both phosphorylated and unphosphorylated RNA Polymerase II (Wen, 1999; Lindstrom, 2001; Lindstrom, 2003; Eissenberg, 2007 and references therein).

The chromo domain motif is a binding site for methylated histone tails. The role of the CHD1 chromo domain in methylated histone binding is controversial. However, recent structural data determined that the double chromo domain of mammalian CHD1 binds methylated H3K4 in vitro (Flanagan, 2005). This study shows that Cdk9 knockdown leads to a loss of chromosomal CHD1. This observation is most easily interpreted as the result of loss of H3K4 methylation that also occurs in Cdk9 knockdown chromosomes. Thus, the finding reported in this study lends support to the in vitro binding data and strongly suggests that the chromo domain-methylated histone interaction plays a dominant role in targeting CHD1 to active chromatin in vivo (Eissenberg, 2007).

The observation that both H3K4 and H3K36 methylation are significantly reduced in Cdk9 knockdown chromosomes suggests a linkage between phosphorylation of the CTD and histone methylation at transcribed genes. In this respect, Cdk9 subsumes activities found in yeast Bur1/Bur2 and yeast Ctk1. Since no significant difference was observed in ASH1 protein levels on Cdk9 knockdown chromosomes, a model is favored in which Cdk9-dependent RNA Polymerase II elongation plays a mechanistic role in H3 tail methylation. In this model, RNA Polymerase II passage destabilizes histone-DNA contacts, making the histones better substrates for efficient methylation. Reduced CTD phosphorylation would lead to reduced rates of RNA Polymerase II transcription genome-wide, resulting in reduced efficiency of histone tail methylation. While the mechanism connecting CTD phosphorylation to RNA Polymerase II elongation rate is likely to be complex in vivo, the observation that reduced CTD phosphorylation is associated with reduced dELL binding suggests that loss of dELL association could be a contributing factor (Eissenberg, 2007).

Mutation in Ash1 in Drosophila results in loss of all detectable H3K4 methylation, but has no effect on H3K36 methylation. This is consistent with independent mechanisms for these two chromatin modifications. A Polymerase II passage model provides a simple mechanism to account for similar effects on both modifications based on substrate availability (Eissenberg, 2007).

Mutational analysis of an RNA polymerase II elongation factor in Drosophila melanogaster

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The ELL family of proteins function in vitro as elongation factors for RNA polymerase II. Deletion studies have defined domains in mammalian ELL required for transcription elongation activity and RNA polymerase binding in vitro, for transformation of cultured cells when overexpressed, and for leukemogenesis and cell proliferation as part of a leukemic fusion protein. The goal of this study was to identify domains required for chromosome targeting and viability in the unique Drosophila ELL (dELL) protein. This study shows that an N-terminal domain of dELL is necessary and sufficient for targeting to transcriptionally active puff sites in chromatin, supporting a role for this domain in recruiting dELL to elongating RNA polymerase II. A central domain of dELL is required for rapid mobilization of ELL during the heat shock response, suggesting a regulatory function for this domain. Unexpectedly, transgenic dELL in which the N-terminal chromosome binding domain is deleted can complement the recessive lethality of mutations in ELL, suggesting that Drosophila ELL has an essential activity in development distinct from its role as an RNA polymerase II elongation factor (Gerber, 2005).

In Drosophila, heat shock results in a dramatic reduction in developmental gene transcription, accompanied by intense transcriptional activation of heat shock genes and the recruitment of Pol II and a number of Pol II elongation factors, to heat shock puff sites in polytene chromosomes. Endogenous dELL quickly disappears from most chromosomal sites after heat shock, and it is rapidly recruited together with phosphorylated Pol II to major heat shock puffs (Gerber, 2001). This mobilization also occurs with the dELL(δ760-1059) mutant but, surprisingly, not with the dELL(δ190-759) mutant, which remains associated with developmental loci after Pol II has largely been lost from these sites. This suggests that while the interaction between dELL and Pol II is dependent upon the N-terminal domain, it is also regulated by an internal domain of dELL. One explanation for this result may be that the internal domain of dELL regulates binding of dELL to transcription factors bound at a variety of developmental genes, and this interaction is disrupted upon heat shock. Thus, the dELL(δ190-759) protein, lacking this regulatory domain, remains bound to developmental loci after heat shock even as Pol II is lost from these loci (Gerber, 2005).

The C-terminal occludin homology domain of mammalian ELL (corresponding to amino acids 839 to 928 in dELL) has been implicated in a number of interactions with potential physiological significance. It is the most highly conserved region of the dELL primary sequence, and it is necessary and sufficient for oncogenic activity in the context of the MLL-ELL fusion protein (DiMartino, 2000; Luo, 2001). It has also been implicated in binding the tumor suppressor p53 (Shinobo, 1999; Wiederschain, 2003) and in the regulation of cell growth and proliferation (Johnstone, 2001). This study shows that this domain is dispensable for chromosome binding and for recruitment of dELL to heat shock loci. Nevertheless, deletion of this domain results in proteins with little or no ability to complement the recessive lethality of hypomorphic Su(Tpl) alleles. While the exact mechanism of the essential requirement for the ELL C-terminal domain remains unknown, it may be significant that deletions of (and point mutations in) the C-terminal domain were uncovered in four separate screens as dominant suppressors of Ras pathway activation. Perhaps this domain provides a mechanistic link between Ras signaling and transcription (Gerber, 2005).

In light of the requirement for the N-terminal domain for dELL chromosome binding, it was unanticipated that an N-terminal domain deletion could complement the recessive lethality of Su(Tpl) mutations. One obvious interpretation of this result is that the recessive lethality of Su(Tpl) mutations is not due to defects in transcription elongation for one or more essential genes but that the essential requirement for dELL in development lies in a distinct dELL-dependent pathway. dELL is one of several elongation factor homologues in Drosophila, and it is possible that the role of dELL in transcription elongation is functionally redundant to these other factors (Gerber, 2005).

Another possibility is that N-terminally truncated dELL functionally complements the Su(Tpl)^S-192 mutant protein, which is present in both complementation assays and which has an intact N-terminal domain but carries missense mutations in the C-terminal domain. This mechanism would suggest that multiple dELL molecules participate in functional protein complexes. Nevertheless, such complexes are presumably extrachromosomal, since the N-terminally truncated dELL does not bind chromosomes even in the presence of wild-type dELL. Future studies aimed at defining the biochemistry of ELL-Pol II complexes should help sort out the mechanism for this genetic interaction (Gerber, 2005).

Drosophila ELL is associated with actively elongating RNA polymerase II on transcriptionally active sites in vivo

Several factors have been biochemically characterized based on their ability to increase the overall rate of transcription elongation catalyzed by the multiprotein complex RNA polymerase II (Pol II). Among these, the ELL family of elongation factors has been shown to increase the catalytic rate of transcription elongation in vitro by suppressing transient pausing. Several fundamental biological aspects of this class of elongation factors are not known. The Drosophila homolog (dELL) was cloned in order to test whether ELL family proteins are actually associated with the elongating Pol II in vivo. dELL is a nuclear protein, which, like its mammalian homologs, can increase the catalytic rate of transcription elongation by Pol II in vitro. Interestingly, it was found that dELL co-localizes extensively with the phosphorylated, actively elongating form of Pol II at transcriptionally active sites on Drosophila polytene chromosomes. Furthermore, dELL is relocalized from a widespread distribution pattern on polytenes under normal conditions to very few transcriptionally active puff sites upon heat shock. This observation indicates a dynamic pattern of localization of dELL in cells, which is a predicted characteristic of a Pol II general elongation factor. It was also demonstrated that dELL physically interacts with Pol II. These results strongly suggest that dELL functions with elongating RNA polymerase II in vivo (Gerber, 2001. Full text of article).

Search PubMed for articles about Drosophila ELL

Banks, C. A., et al. (2007). Identification and Characterization of a Schizosaccharomyces pombe RNA polymerase II elongation factor with similarity to the metazoan transcription factor ELL. J. Biol. Chem. 282: 5761-5769. PubMed ID: 17150956

DiMartino, J. F., T. Miller, P. M. Ayton, T. Landewe, J. L. Hess, M. L. Cleary and A. Shilatifard (2000). A carboxy-terminal domain of ELL is required and sufficient for immortalization of myeloid progenitors by MLL-ELL. Blood 96: 3887-3893. PubMed ID: 11090074

Eissenberg, J. C., et al. (2002). dELL is an essential RNA polymerase II elongation factor with a general role in development. Proc. Natl. Acad. Sci. 99: 9894-9899. PubMed ID: 12096188

Eissenberg, J. C., Shilatifard, A., Dorokhov, N., Michener, D. E.. (2007). Cdk9 is an essential kinase in Drosophila that is required for heat shock gene expression, histone methylation and elongation factor recruitment. Mol. Genet. Genomics. 277(2): 101-14. PubMed ID: 17001490

Flanagan, J. F., et al. (2005). Double chromodomains cooperate to recognize the methylated histone H3 tail. Nature 438(7071): 1181-5. PubMed ID: 16372014

Gerber, M., et al. (2001). Drosophila ELL is associated with actively elongating RNA polymerase II on transcriptionally active sites in vivo. EMBO J. 20(21): 6104-14. PubMed ID: 11689450

Gerber, M., et al. (2004). In vivo requirement of the RNA polymerase II elongation factor elongin A for proper gene expression and development. Mol. Cell Biol. 24: 9911-9919. PubMed ID: 15509793

Gerber, M. A., Shilatifard, A. and Eissenberg, J. C. (2005). Mutational analysis of an RNA polymerase II elongation factor in Drosophila melanogaster. Mol. Cell Biol. 25(17): 7803-11. PubMed ID: 16107725

Johnstone, R. W., et al. (2001). Functional analysis of the leukemia protein ELL: evidence for a role in the regulation of cell growth and survival. Mol. Cell. Biol. 21: 1672-1681. PubMed ID: 11238904

Kong, S. E., et al. (2005) ELL-associated factors 1 and 2 are positive regulators of RNA polymerase II elongation factor ELL. Proc. Natl. Acad. Sci. 102: 10094-10098. PubMed ID: 16006523

Lindstrom, D. L. and Hartzog, G. A. (2001). Genetic interactions of Spt4-Spt5 and TFIIS with the RNA polymerase II CTD and CTD modifying enzymes in Saccharomyces cerevisiae. Genetics 159: 487-497. PubMed ID: 11606527

Lindstrom, D. L., et al. (2003). Dual roles for Spt5 in pre-mRNA processing and transcription elongation revealed by identification of Spt5-associated proteins. Mol. Cell Biol. 23: 1368-1378. PubMed ID: 12556496

Luo, R. T., et al. (2001). The elongation domain of ELL is dispensable but its ELL-associated factor 1 interaction domain is essential for MLL-ELL-induced leukemogenesis. Mol. Cell. Biol. 21: 5678-5687. PubMed ID: 11463848

Muse, G. W., et al. (2007). RNA polymerase is poised for activation across the genome. Nat. Genet. 39: 1507-1511. PubMed ID: 17994021

Shilatifard, A., Conaway, R. C. and Conaway, J. W. (2003). The RNA polymerase II elongation complex. Annu. Rev. Biochem. 72: 693-715. PubMed ID: 12676794

Shinobu, N., et al. (1999). Physical interaction and functional antagonism between the RNA polymerase II elongation factor ELL and p53. J. Biol. Chem. 274: 17003-17010. PubMed ID: 10358050

Simone, F., et al. (2001). EAF1, a novel ELL-associated factor that is delocalized by expression of the MLL-ELL fusion protein. Blood 98: 201-209. PubMed ID: 11418481

Simone, F., et al. (2003). ELL-associated factor 2 (EAF2), a functional homolog of EAF1 with alternative ELL binding properties. Blood 101: 2355-2362. PubMed ID: 12446457

Smith, E. R., Winter, B., Eissenberg, J. C. and Shilatifard, A. (2008). Regulation of the transcriptional activity of poised RNA polymerase II by the elongation factor ELL. Proc. Natl. Acad. Sci. 105(25): 8575-9. PubMed ID: 18562276

Wen, Y. and Shatkin, A. J. (1999). Transcription elongation factor hSPT5 stimulates mRNA capping. Genes Dev 13: 1774-1779. PubMed ID: 10421630

Wiederschain, D., et al. (2003). Molecular basis of p53 functional inactivation by the leukemic protein MLL-ELL. Mol. Cell. Biol. 23: 4230-4246. PubMed ID: 12773566

Zeitlinger, J., et al. (2007). RNA polymerase stalling at developmental control genes in the Drosophila melanogaster embryo. Nat. Genet. 39: 1512-1516. PubMed ID: 17994019

date revised: 3 September 2009

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