The intranuclear position of many genes has been correlated with their activity state, suggesting that migration to functional subcompartments may influence gene expression. Indeed, nascent RNA production and RNA polymerase II seem to be localized into discrete foci or 'transcription factories'. Current estimates from cultured cells indicate that multiple genes could occupy the same factory, although this has not yet been observed. Here we show that, during transcription in vivo, distal genes colocalize to the same transcription factory at high frequencies. Active genes are dynamically organized into shared nuclear subcompartments, and movement into or out of these factories results in activation or abatement of transcription. Thus, rather than recruiting and assembling transcription complexes, active genes migrate to preassembled transcription sites.

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Active genes dynamically colocalize to shared sites of ongoing transcription.

The superfamily of tripartite motif-containing (TRIM) proteins is conserved throughout the metazoan kingdom and has expanded rapidly during vertebrate evolution; there are now more than 60 TRIM proteins known in humans and mice. Many TRIM proteins are induced by type I and type II interferons, which are crucial for many aspects of resistance to pathogens, and several are known to be required for the restriction of infection by lentiviruses. In this Review, we describe recent data that reveal broader antiviral and antimicrobial activities of TRIM proteins and discuss their involvement in the regulation of pathogen-recognition and transcriptional pathways in host defence.

TRIM family proteins and their emerging roles in innate immunity

The yeast Sir2 protein mediates chromatin silencing through an intrinsic NAD-dependent histone deacetylase activity. Sir2 is a conserved protein and was recently shown to regulate lifespan extension both in budding yeast and worms. Here, we show that SIRT1, the human Sir2 homolog, is recruited to the promyelocytic leukemia protein (PML) nuclear bodies of mammalian cells upon overexpression of either PML or oncogenic Ras (Ha-rasV12). SIRT1 binds and deacetylates p53, a component of PML nuclear bodies, and it can repress p53-mediated transactivation. Moreover, we show that SIRT1 and p53 co-localize in nuclear bodies upon PML upregulation. When overexpressed in primary mouse embryo fibroblasts (MEFs), SIRT1 antagonizes PML-induced acetylation of p53 and rescues PML-mediated premature cellular senescence. Taken together, our data establish the SIRT1 deacetylase as a novel negative regulator of p53 function capable of modulating cellular senescence.

/pdf/human-sir2-deacetylates-p53-and-antagonizes-pml-p53-induced-7i7js6ycez.pdf

Human SIR2 deacetylates p53 and antagonizes PML/p53-induced cellular senescence

The promyelocytic leukaemia (PML) tumour suppressor protein epitomizes the PML-nuclear body (PML-NB) and is crucially required for the proper assembly of this macromolecular nuclear structure. Unlike other, more specialized subnuclear structures such as Cajal and Polycomb group bodies, PML-NBs are functionally promiscuous and have been implicated in the regulation of diverse cellular functions. PML-NBs are dynamic structures that favour the sequestration and release of proteins, mediate their post-translational modifications and promote specific nuclear events in response to various cellular stresses. Recent data suggest that PML-NBs may be heterogeneous in composition, mobility and function.

Structure, dynamics and functions of promyelocytic leukaemia nuclear bodies

The many faces of histone lysine methylation.

PML is a component of a multiprotein complex, termed nuclear bodies, and the PML protein was originally discovered in patients suffering from acute promyelocytic leukaemia (APL). APL is associated with a reciprocal chromosomal translocation of chromosomes 15 and 17, which results in a fusion protein comprising PML and the retinoic acid receptor alpha. The PML genomic locus is approximately 35 kb and is subdivided into nine exons. A large number of alternative spliced transcripts are synthesized from the PML gene, resulting in a variety of PML proteins ranging in molecular weight from 48-97 kDa. In this review we summarize the data on the known PML isoforms and splice variants and present a new unifying nomenclature. Although, the function/s of the PML variants are unclear, all PML isoforms contain an identical N-terminal region, suggesting that these sequences are indispensable for function, but differ in their C-terminal sequences. The N-terminal region harbours a RING-finger, two B-boxes and a predicted alpha-helical Coiled-Coil domain, that together form the RBCC/TRIM motif found in a large family of proteins. In PML this motif is essential for PML nuclear body formation in vivo and PML-homo and hetero interactions conferring growth suppressor, apoptotic and anti-viral activities. In APL oligomerization mediated by the RBCC/TRIM motif is essential for the transformation potential of the PML-RARalpha fusion protein.

PML protein isoforms and the RBCC/TRIM motif

The Polycomb group (PcG) complex is a chromatin-associated multiprotein complex, involved in the stable repression of homeotic gene activity in Drosophila. Recently, a mammalian PcG complex has been identified with several PcG proteins implicated in the regulation of Hox gene expression. Although the mammalian PcG complex appears analogous to the complex in Drosophila, the molecular mechanisms and functions for the mammalian PcG complex remain unknown. Here we describe a detailed characterization of the human PcG complex in terms of cellular localization and chromosomal association. By using antibodies that specifically recognize three human PcG proteins— RING1, BMI1, and hPc2—we demonstrate in a number of human cell lines that the PcG complex forms a unique discrete nuclear structure that we term PcG bodies. PcG bodies are prominent novel nuclear structures with the larger PcG foci generally localized near the centromeres, as visualized with a kinetochore antibody marker. In both normal fetal and adult fibroblasts, PcG bodies are not randomly dispersed, but appear clustered into defined areas within the nucleus. We show in three different human cell lines that the PcG complex can tightly associate with large pericentromeric heterochromatin regions (1q12) on chromosome 1, and with related pericentromeric sequences on different chromosomes, providing evidence for a mammalian PcG–heterochromatin association. Furthermore, these heterochromatin-bound PcG complexes remain stably associated throughout mitosis, thereby allowing the potential inheritance of the PcG complex through successive cell divisions. We discuss these results in terms of the known function of the PcG complex as a transcriptional repression complex.

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The Human Polycomb Group Complex Associates with Pericentromeric Heterochromatin to Form a Novel Nuclear Domain

The promyelocytic leukemia (PML) protein is aggregated into nuclear bodies that are associated with diverse nuclear processes. Here, we report that the distance between a locus and its nearest PML body correlates with the transcriptional activity and gene density around the locus. Genes on the active X chromosome are more significantly associated with PML bodies than their silenced homologues on the inactive X chromosome. We also found that a histone-encoding gene cluster, which is transcribed only in S-phase, is more strongly associated with PML bodies in S-phase than in G0/G1 phase of the cell cycle. However, visualization of specific RNA transcripts for several genes showed that PML bodies were not themselves sites of transcription for these genes. Furthermore, knock-down of PML bodies by RNA interference did not preferentially change the expression of genes closely associated with PML bodies. We propose that PML bodies form in nuclear compartments of high transcriptional activity, but they do not directly regulate transcription of genes in these compartments.

/pdf/promyelocytic-leukemia-nuclear-bodies-associate-with-42oce8vb11.pdf

Promyelocytic leukemia nuclear bodies associate with transcriptionally active genomic regions.

Promyelocytic leukemia (PML) bodies are nuclear multi-protein domains. The observations that viruses transcribe their genomes adjacent to PML bodies and that nascent RNA accumulates at their periphery suggest that PML bodies function in transcription. We have used immuno-FISH in primary human fibroblasts to determine the 3D spatial organisation of gene-rich and gene-poor chromosomal regions relative to PML bodies. We find a highly non-random association of the gene-rich major histocompatibilty complex (MHC) on chromosome 6 with PML bodies. This association is specific for the centromeric end of the MHC and extends over a genomic region of at least 1.6 megabases. We also show that PML association is maintained when a subsection of this region is integrated into another chromosomal location. This is the first demonstration that PML bodies have specific chromosomal associations and supports a model for PML bodies as part of a functional nuclear compartment.

Carol Shiels

Papers

PML protein isoforms and the RBCC/TRIM motif

The Human Polycomb Group Complex Associates with Pericentromeric Heterochromatin to Form a Novel Nuclear Domain

Promyelocytic leukemia nuclear bodies associate with transcriptionally active genomic regions.

PML bodies associate specifically with the MHC gene cluster in interphase nuclei.

Contiguous Arrays of Satellites 1, 3, and β Form a 1.5-Mb Domain on Chromosome 22p