Interplay between PML NBs and HIRA for H3.3 deposition on transcriptionally active interferon-stimulated genes

Promyelocytic Leukemia Nuclear Bodies (PML NBs) are nuclear membrane-less organelles physically associated with chromatin underscoring their crucial role in genome function. The H3.3 histone chaperone complex HIRA accumulates in PML NBs upon senescence, viral infection or IFN-I treatment in primary cells. Yet, the molecular mechanisms of this partitioning and its function in regulating histone dynamics have remained elusive. Here, by using specific siRNAs and protein Affimers, we identify intermolecular SUMO-SIM interactions as an essential mechanism for HIRA recruitment in PML NBs. In addition, we demonstrate that HIRA localization in the nuclear bodies is intimately linked to the presence of a soluble pool of H3.3-H4 dimers inside PML NBs, that is not found in cancer cells. Transcription inhibition prevents HIRA accumulation in PML NBs underscoring the importance of transcriptional activity to drive HIRA through PML NBs. Finally, in the context of inflammatory responses, HIRA and PML are necessary for the prolonged H3.3 deposition at the transcriptional end sites of interferon-stimulated genes (ISGs), well beyond the peak of transcription. We thus propose that HIRA partitioning in PML NBs is essential to regulate H3.3 deposition on transcriptionally active regions.

CCT8 recovers WTp53-suppressed cell cycle evolution and EMT to promote colorectal cancer progression

LIM and SH3 protein 1 (LASP1) is a metastasis-related protein reported to enhance tumor progression in colorectal cancer (CRC). However, the underlying mechanism is still elusive. The chaperonin protein containing TCP1 (CCT) is a cellular molecular chaperone complex, which is necessary for the correct folding of many proteins. It contains eight subunits, CCT1-8. CCT8 is overexpressed in many cancers, however, studies on CCT8 are limited and its role on CRC development and progression remains elusive. In this study, we confirmed that CCT8 and LASP1 can interact with each other and express positively in CRC cells. CCT8 could recover the ability of LASP1 to promote the invasion of CRC; CCT8 could significantly promote the proliferation, invasion, and metastasis of colorectal cells in vivo and in vitro. Mechanically, CCT8 inhibited the entry of WTp53 into the nucleus, and there was a negative correlation between the expression of CCT8 and the nuclear expression of WTp53 in clinical colorectal tissues. CCT8 promoted the cell cycle evolution and EMT progression of CRC by inhibiting the entry of WTp53 into the nucleus. Clinically, CCT8 was highly expressed in CRC. More importantly, the overall survival of CRC patients with high expression of CCT8 was worse than that of patients with low expression of CCT8. These findings indicate that as LASP1-modulated proteins, CCT8 plays a key role in promoting the progression of colorectal cancer, which provides a potential target for clinical intervention in patients with colorectal cancer.

NASP maintains histone H3–H4 homeostasis through two distinct H3 binding modes

Histone chaperones regulate all aspects of histone metabolism. NASP is a major histone chaperone for H3–H4 dimers critical for preventing histone degradation.Here, we identify two distinct histone binding modes of NASP and reveal how they cooperate to ensure histone H3–H4 supply. We determine the structures of a sNASP dimer, a complex of sNASP with an H3 α3 peptide, and the sNASP–H3–H4–ASF1b co-chaperone complex.

DAXX-ATRX-H3.3 Regulation of p53 Chromatin Binding and DNA Damage Response

DAXX and ATRX are tumor suppressor proteins that form a histone H3.3 chaperone complex and are frequently mutated in cancers with the alternative lengthening of telomeres (ALT), such as pediatric glioblastoma. Rapid loss of function of either DAXX or ATRX are not by themselves sufficient to induce the ALT phenotype. However, cells lacking DAXX or ATRX can be readily selected for ALT-like features. Here, we show that DAXX and ATRX null glioblastoma cells with ALT-like features have defects in p53 chromatin binding and DNA damage response regulation. RNA-seq analysis of DAXX or ATRX null U87 glioblastoma cells with ALT-like features revealed that p53 pathway is among perturbed. ALT-selected DAXX and ATRX-null cells had aberrant response to DNA damaging agent etoposide. Both DAXX and ATRX-null ALT cells showed a loss of p53 binding at a subset of response elements. Complementation of DAXX null cells with a wild-type DAXX transgene rescued p53 binding and transcription, while the tumor associated mutation L130R that disrupts ATRX binding was incapable of rescuing p53 chromatin binding. We show that histone H3.3 binding is reduced in DAXX-null cells especially at subtelomeric p53 binding sites and telomere repeats. These findings indicate that DAXX and ATRX function to enable p53 chromatin binding through modulation of histone H3.3 binding, especially at sub-telomeric sites.

The Hsp70–Hsp90 go-between Hop/Stip1/Sti1 is a proteostatic switch and may be a drug target in cancer and neurodegeneration

The Hsp70 and Hsp90 molecular chaperone systems are critical regulators of protein homeostasis (proteostasis) in eukaryotes under normal and stressed conditions. The Hsp70 and Hsp90 systems physically and functionally interact to ensure cellular proteostasis. Co-chaperones interact with Hsp70 and Hsp90 to regulate and to promote their molecular chaperone functions. Mammalian Hop, also called Stip1, and its budding yeast ortholog Sti1 are eukaryote-specific co-chaperones, which have been thought to be essential for substrate (“client”) transfer from Hsp70 to Hsp90. Substrate transfer is facilitated by the ability of Hop to interact simultaneously with Hsp70 and Hsp90 as part of a ternary complex. Intriguingly, in prokaryotes, which lack a Hop ortholog, the Hsp70 and Hsp90 orthologs interact directly. Recent evidence shows that eukaryotic Hsp70 and Hsp90 can also form a prokaryote-like binary chaperone complex in the absence of Hop, and that this binary complex displays enhanced protein folding and anti-aggregation activities. The canonical Hsp70-Hop-Hsp90 ternary chaperone complex is essential for optimal maturation and stability of a small subset of clients, including the glucocorticoid receptor, the tyrosine kinase v-Src, and the 26S/30S proteasome. Whereas many cancers have increased levels of Hop, the levels of Hop decrease in the aging human brain. Since Hop is not essential in all eukaryotic cells and organisms, tuning Hop levels or activity might be beneficial for the treatment of cancer and neurodegeneration.

Melatonin potentiates the cytotoxic effect of Neratinib in HER2+ breast cancer through promoting endocytosis and lysosomal degradation of HER2

Complete blockade of the HER2 protein itself and HER signaling network is critical to achieving effective HER2-targeted therapies. Despite the success of HER2-targeted therapies, the diseases will relapse in a significant fraction of patients with HER2+ breast cancers. How to improve the therapeutic efficacy of existing HER2-targeted agents remains an unmet clinical need. Here, we uncover a role of Melatonin in diminishing HER2-mediated signaling by destruction of HER2 protein. Mechanistically, Melatonin treatment attenuated the protective effect of the HSP90 chaperone complex on its client protein HER2, triggering ubiquitylation and subsequent endocytic lysosomal degradation of HER2. The inhibitory effect of Melatonin on HER2 signaling substantially enhanced the cytotoxic effects of the pan-HER inhibitor Neratinib in HER2+ breast cancer cells. Lastly, we demonstrate that dual inhibition of HER2 by combined use of Melatonin and Neratinib effectively blocked the growth of HER2+ breast tumor xenografts in vivo. Our findings shed light on the potential use of Melatonin in a novel dual HER2 blockade strategy for HER2+ breast cancer treatment.

Δ133p53β isoform pro-invasive activity is regulated through an aggregation-dependent mechanism in cancer cells

The p53 isoform, Δ133p53β, is critical in promoting cancer. Here we report that Δ133p53β activity is regulated through an aggregation-dependent mechanism. Δ133p53β aggregates were observed in cancer cells and tumour biopsies. The Δ133p53β aggregation depends on association with interacting partners including p63 family members or the CCT chaperone complex. Depletion of the CCT complex promotes accumulation of Δ133p53β aggregates and loss of Δ133p53β dependent cancer cell invasion. In contrast, association with p63 family members recruits Δ133p53β from aggregates increasing its intracellular mobility. Our study reveals novel mechanisms of cancer progression for p53 isoforms which are regulated through sequestration in aggregates and recruitment upon association with specific partners like p63 isoforms or CCT chaperone complex, that critically influence cancer cell features like EMT, migration and invasion.