The nature of the structural changes induced by histone acetylation at the different levels of chromatin organization has been very elusive. At the histone level, it has been proposed on several occasions that acetylation may induce an ␣-helical conformation of their acetylated N-terminal domains (tails). In an attempt to provide experimental support for this hypothesis, we have purified and characterized the tail of histone H4 in its native and mono-, di-, tri-, and tetra-acetylated form. The circular dichroism analysis of these peptides shows conclusively that acetylation does increase their ␣-helical content. Furthermore, the same spectroscopic analysis shows that this is also true for both the acetylated nucleosome core particle and the whole histone octamer in solution. In contrast to the native tails in which the ␣-helical organization appears to be dependent upon interaction of these histone regions with DNA, the acetylated tails show an increase in ␣-helical content that does not depend on such an interaction.

Despite the fact that histone H2A ubiquitination affects about 10-15% of this histone in most eukaryotic cells, histone ubiquitination is among one of the less-well-characterized post-translational histone modifications. Nevertheless, some important observations have been made in recent years. Whilst several enzymes had been known to ubiquitinate histones in vitro, recent studies in yeast have led to the unequivocal identification of the enzyme responsible for this post-translational modification in this organism. A strong functional co-relation to meiosis and spermiogenesis has also now been well documented, although its participation in other functional aspects of chromatin metabolism, such as transcription or DNA repair, still remains rather speculative and controversial. Because of its nature, histone ubiquitination represents the most bulky structural change to histones and as such it would be expected to exert an important effect on chromatin structure. Past and recent structural studies, however, indicate a surprising lack of effect of (H2A/H2B) ubiquitination on nucleosome architecture and of uH2A on chromatin folding. These results suggest that this modification may serve as a signal for recognition by functionally relevant trans-acting factors and/or operate synergistically in conjunction with other post-translational modifications such as for instance acetylation.

We define stress-induced adaptive survival pathways linking autophagy with the molecular chaperone clusterin (CLU) that function to promote anticancer treatment resistance. During treatment stress, CLU co-localizes with LC3 via an LIR-binding sequence within autophagosome membranes, functioning to facilitate LC3–Atg3 heterocomplex stability and LC3 lipidation, and thereby enhance autophagosome biogenesis and autophagy activation. Stress-induced autophagy is attenuated with CLU silencing in CLU−/− mice and human prostate cancer cells. CLU-enhanced cell survival occurs via autophagy-dependent pathways, and is reduced following autophagy inhibition. Combining CLU inhibition with anticancer treatments attenuates autophagy activation, increases apoptosis and reduces prostate cancer growth. This study defines a novel adaptor protein function for CLU under stress conditions, and highlights how co-targeting CLU and autophagy can amplify proteotoxic stress to delay cancer progression.

The folding ability of chromatin fractions containing approximately identical nucleosome numbers and the same linker histone composition, but with different extents of core histone acetylation, were analyzed by analytical ultracentrifugation. It was found that the acetylated fractions consistently exhibited a relatively small but significantly lower extent of compaction than that of their native nonacetylated counterparts. This was regardless of the extent of the size distribution heterogeneity of the fractions analyzed. Furthermore the acetylated chromatin fibers exhibited an enhanced solubility in both NaCl and MgCl 2 , which is neither the result of a differential binding affinity of the linker histones to chromatin nor of an alteration in the relative amounts of the histone H1 variants.The possible implications of histone acetylation for eukaryotic gene transcription were recognized almost from the discovery of this post-translational histone modification more than 35 years ago (1, 2). The finding 15 years later that n-butyrate could increase the levels of histone acetylation in HeLa and Friend erythroleukemic cells (3) represented an important landmark for the structural studies designed to elucidate the structural implications of acetylation because it allowed for the production of the large amounts of material that are usually required for this kind of analyses. However, despite extensive experimental effort in the following years, no significant differences could be found either at the level of the nucleosome chromatin subunit (4,5) or at the level of chromatin fiber folding (6, 7).The discovery that histone acetyltransferases (8) are an integral part of the basal transcription complexes has rekindled interest in histone acetylation as an important factor in the modulation of eukaryotic gene expression (9). Histone acetylation has been linked to cancer (10 -13), and histone deacetylase inhibitors are being used now for the treatment of certain cancer types (14).Despite this, the structural implications of histone acetylation in mediating eukaryotic transcription remain to be established. Thus, although the functional implications seem clear, the mechanisms remain to be unraveled. Although the current coding hypothesis (15) would explain the localized "short range" effects, an example of a transacting factor that requires histone acetylation for its interaction with the chromatin template has not yet been identified. Furthermore the fact that acetylation can occur over long stretches (several kilobases) of DNA (16) ("long range" effect) argues against the coding hypothesis being the only structural role for histone acetylation.In the present paper we have revisited some of the earlier structural analyses using well defined chromatin fractions that differ only in their extent of core histone acetylation. Although our results generally agree with most of the earlier data, they also underscore the structural differences and constraints that may be important for understanding the mechanisms by which histone a...

MacroH2A (mH2A) is one of the most recently identified members of the heteromorphous histone variant family. It is unique among the members of this group because it contains an unusually large non-histone C-terminal end, from where its name derives, and appears to be restricted to subphylum vertebrata. Although a concerted effort has been carried out in order to characterize the physiological relevance of mH2A, little is known in comparison about the structural importance of the molecule. Elucidating the biophysical and conformational proprieties of mH2A in chromatin may provide clues into the links between this histone variant and its unique function(s). In this paper, we look first at the heterogeneous tissue-specific distribution of this protein in different vertebrate classes. This is followed by a structural comparison between mH2A and H2A protein and by the characterization of the nucleosome core particles with which these histone subtypes are associated. We find that the highly alpha-helical C-terminus of mH2A confers an asymmetric conformation to nucleosomes and that this variant is tightly bound to chromatin fragments in a way that does not depend on the overall extent of acetylation of the other core histones.