Ramin Shiekhattar

Breast and ovarian cancer susceptibility genes BRCA1 and PALB2 have enigmatic roles in cellular growth and mammalian development. While these genes are essential for growth during early developmental programs, inactivation later in adulthood results in increased growth and formation of tumors, leading to their designation as tumor suppressors. We performed genome‐wide analysis assessing their chromatin residence and gene expression responsiveness using high‐throughput sequencing in breast epithelial cells. We found an intimate association between BRCA1 and PALB2 chromatin residence and genes displaying high transcriptional activity. Moreover, our experiments revealed a critical role for BRCA1 and, to a smaller degree, PALB2 in transcriptional responsiveness to NF‐κB, a crucial mediator of growth and inflammatory response during development and cancer. Importantly, we also uncovered a vital role for BRCA1 and PALB2 in response to retinoic acid (RA), a growth inhibitory signal in breast cancer cells, which may constitute the basis for their tumor suppressor activity. Taken together, our results highlight an important role for these breast cancer proteins in the regulation of diverse growth regulatory pathways. ![][1] High‐throughput sequencing reveals chromatin‐binding sites of the breast cancer tumor suppressors BRCA1 and PALB2 and suggests transcription co‐regulatory roles at a subset of genes, including NF‐κB targets and retinoic acid‐responsive genes. EMBO J (2014) 33, 890–905 [1]: /embed/graphic-1.gif

Cell death and differentiation is a monthly research journal focused on the exciting field of programmed cell death and apoptosis. It provides a single accessible source of information for both scientists and clinicians, keeping them up-to-date with advances in the field. It encompasses programmed cell death, cell death induced by toxic agents, differentiation and the interrelation of these with cell proliferation.

A new study identifies the long noncoding RNA Pint as a regulator of cellular proliferation and a target of the p53 pathway. See related Research, http://genomebiology.com/2013/14/9/R104

Long noncoding RNAs are emerging as a critical component of transcriptional activation. Over the past few years, there has been a number of examples where long noncoding RNAs expressed from distal sites exert their activating functions on neighbouring protein‐coding genes. A new study in The EMBO Journal (Lu et al , 2013) now extends this paradigm by revealing a role for the transcription factor Yin Yang 1 (YY1) in the regulation of the expression of long noncoding RNAs (termed Yams) during myogenesis. Importantly, the authors demonstrate a functional role for Yam‐1 in muscle differentiation through activation of a neighbouring micoRNA‐715 leading to downregulation of Wnt7b. There is converging evidence that distal regulatory sites commonly known as ‘enhancers’ are transcribed in a stimulus‐dependent manner as long noncoding RNAs (known as enhancer RNAs, eRNAs) and such eRNAs contribute to transcriptional activation of their neighbouring genes (De Santa et al , 2010; Kim et al , 2010; Hah et al , 2013; Lam et al , 2013; Li et al , 2013; Melo et al , 2013). Concomitantly, studies of long noncoding RNAs located in intergenic sites (termed lincRNAs, large intergenic noncoding RNAs) have uncovered a role for a class of lincRNAs (named noncoding RNA‐activating, ncRNA‐a) having enhancer‐like functions (Orom et al , 2010; Wang et al , 2011). These activating lincRNAs are enriched with transcription factor binding sites at their promoters and display responsiveness to differentiating stimuli (Orom et al, …

A class of long non-coding RNA (lncRNA) with enhancer-like activity is found to associate with the co-activator complex Mediator and promote its genomic association and enzymatic activity; together with Mediator, the lncRNAs also help to maintain the chromosomal architecture of active regulatory elements. Long non-coding RNAs (lncRNAs) can both repress and activate gene expression. Here, a class of lncRNAs with enhancer-like activity is found to associate with the translational co-activator complex Mediator. Termed ncRNA-activating (ncRNA-a), these molecules promote the genomic association and enzymatic activity of Mediator, and acting together with Mediator, they also help to maintain the chromosomal architecture of active regulatory elements. Importantly, Mediator complexes containing disease-linked mutant MED12 proteins fail to associate with ncRNA-a. The MED12 gene encodes a Mediator complex subunit, and MED12 mutations have been linked to FG syndrome, a rare genetic disorder with symptoms including intellectual disability. This work suggests that the loss of Mediator–ncRNA-a interactions might be a possible contributing factor in such developmental diseases. Recent advances in genomic research have revealed the existence of a large number of transcripts devoid of protein-coding potential in multiple organisms1,2,3,4,5,6,7,8. Although the functional role for long non-coding RNAs (lncRNAs) has been best defined in epigenetic phenomena such as X-chromosome inactivation and imprinting, different classes of lncRNAs may have varied biological functions8,9,10,11,12,13. We and others have identified a class of lncRNAs, termed ncRNA-activating (ncRNA-a), that function to activate their neighbouring genes using a cis-mediated mechanism5,14,15,16. To define the precise mode by which such enhancer-like RNAs function, we depleted factors with known roles in transcriptional activation and assessed their role in RNA-dependent activation. Here we report that depletion of the components of the co-activator complex, Mediator, specifically and potently diminished the ncRNA-induced activation of transcription in a heterologous reporter assay using human HEK293 cells. In vivo, Mediator is recruited to ncRNA-a target genes and regulates their expression. We show that ncRNA-a interact with Mediator to regulate its chromatin localization and kinase activity towards histone H3 serine 10. The Mediator complex harbouring disease-17,18 displays diminished ability to associate with activating ncRNAs. Chromosome conformation capture confirmed the presence of DNA looping between the ncRNA-a loci and its targets. Importantly, depletion of Mediator subunits or ncRNA-a reduced the chromatin looping between the two loci. Our results identify the human Mediator complex as the transducer of activating ncRNAs and highlight the importance of Mediator and activating ncRNA association in human disease.

Initiation of transcription of RNA polymerase II (RNAPII)-dependent genes requires the participation of a host of basal transcription factors. Among genes requiring RNAPII for transcription, small nuclear RNAs (snRNAs) display a further requirement for a factor known as snRNA-activating protein complex (SNAPc). The scope of the biological function of SNAPc and its requirement for transcription of protein-coding genes has not been elucidated. To determine the genome-wide occupancy of SNAPc, we performed chromatin immunoprecipitation followed by high-throughput sequencing using antibodies against SNAPC4 and SNAPC1 subunits. Interestingly, while SNAPC4 occupancy was limited to snRNA genes, SNAPC1 chromatin residence extended beyond snRNA genes to include a large number of transcriptionally active protein-coding genes. Notably, SNAPC1 occupancy on highly active genes mirrored that of elongating RNAPII extending through the bodies and 3' ends of protein-coding genes. Inhibition of transcriptional elongation resulted in the loss of SNAPC1 from the 3' ends of genes, reflecting a functional association between SNAPC1 and elongating RNAPII. Importantly, while depletion of SNAPC1 had a small effect on basal transcription, it diminished the transcriptional responsiveness of a large number of genes to two distinct extracellular stimuli, epidermal growth factor (EGF) and retinoic acid (RA). These results highlight a role for SNAPC1 as a general transcriptional coactivator that functions through elongating RNAPII.

Small GTPases share a biochemical mechanism and act as binary molecular switches. One important function of small GTPases in the cell is nucleocytoplasmic transport of both proteins and RNA. Here, we show the stable association of human GPN1 and GPN3, small GTPases related to Ran, with RNA polymerase II (RNAPII) isolated from either the cytoplasmic or nuclear fraction. GPN1 and GPN3 directly interact with RNAPII subunit 7 (RPB7)/RPB4 and the C-terminal domain (CTD) of RNAPII. Depletion of GPN1 or GPN3 using small interfering RNAs led to decreased RNAPII levels in the nucleus and an accumulation of this enzyme in the cytoplasm of human cells. Furthermore, isolation of a GPN1/GPN3/RNAPII complex from stable cell lines expressing a dominant negative GPN1 harboring mutations in the GTP-binding pocket demonstrated a role for these proteins in nuclear import of RNAPII. Thus, GPN1/GPN3 define a new family of small GTPases that are specialized for the transport of RNA polymerase II into the nucleus.