Chromosomal instability as a driver of cancer progression.

Chromosomal instability (CIN) refers to an increased propensity of cells to acquire structural and numerical chromosomal abnormalities during cell division, which contributes to tumour genetic heterogeneity. CIN has long been recognized as a hallmark of cancer, and evidence over the past decade has strongly linked CIN to tumour evolution, metastasis, immune evasion and treatment resistance. Until recently, the mechanisms by which CIN propels cancer progression have remained elusive. Beyond the generation of genomic copy number heterogeneity, recent work has unveiled additional tumour-promoting consequences of abnormal chromosome segregation. These mechanisms include complex chromosomal rearrangements, epigenetic reprogramming and the induction of cancer cell-intrinsic inflammation, emphasizing the multifaceted role of CIN in cancer.

Epigenetic dysregulation from chromosomal transit in micronuclei.

Chromosomal instability (CIN) and epigenetic alterations are characteristics of advanced and metastatic cancers1-4, but whether they are mechanistically linked is unknown. Here we show that missegregation of mitotic chromosomes, their sequestration in micronuclei5,6 and subsequent rupture of the micronuclear envelope7 profoundly disrupt normal histone post-translational modifications (PTMs), a phenomenon conserved across humans and mice, as well as in cancer and non-transformed cells. Some of the changes in histone PTMs occur because of the rupture of the micronuclear envelope, whereas others are inherited from mitotic abnormalities before the micronucleus is formed. Using orthogonal approaches, we demonstrate that micronuclei exhibit extensive differences in chromatin accessibility, with a strong positional bias between promoters and distal or intergenic regions, in line with observed redistributions of histone PTMs. Inducing CIN causes widespread epigenetic dysregulation, and chromosomes that transit in micronuclei experience heritable abnormalities in their accessibility long after they have been reincorporated into the primary nucleus. Thus, as well as altering genomic copy number, CIN promotes epigenetic reprogramming and heterogeneity in cancer.

Engineered Coiled-Coil Protein for Delivery of Inverse Agonist for Osteoarthritis.

Osteoarthritis (OA) results from degenerative and abnormal function of joints, with localized biochemistry playing a critical role in its onset and progression. As high levels of all- trans retinoic acid (ATRA) in synovial fluid have been identified as a contributive factor to OA, the synthesis of de novo antagonists for retinoic acid receptors (RARs) has been exploited to interrupt the mechanism of ATRA action. BMS493, a pan-RAR inverse agonist, has been reported as an effective inhibitor of ATRA signaling pathway; however, it is unstable and rapidly degrades under physiological conditions. We employed an engineered cartilage oligomeric matrix protein coiled-coil (CccS) protein for the encapsulation, protection, and delivery of BMS493. In this study, we determine the binding affinity of CccS to BMS493 and the stimulator, ATRA, via competitive binding assay, in which ATRA exhibits approximately 5-fold superior association with CccS than BMS493. Interrogation of the structure of CccS indicates that ATRA causes about 10% loss in helicity, while BMS493 did not impact the structure. Furthermore, CccS self-assembles into nanofibers when bound to BMS493 or ATRA as expected, displaying 11-15 nm in diameter. Treatment of human articular chondrocytes in vitro reveals that CccS·BMS493 demonstrates a marked improvement in efficacy in reducing the mRNA levels of matrix metalloproteinase-13 (MMP-13), one of the main proteases responsible for the degradation of the extracellular cartilage matrix compared to BMS493 alone in the presence of ATRA, interleukin-1 beta (IL-1ß), or IL-1 ß together with ATRA. These results support the feasibility of utilizing coiled-coil proteins as drug delivery vehicles for compounds of relatively limited bioavailability for the potential treatment of OA.

Micronuclear collapse from oxidative damage.

Chromosome-containing micronuclei are a hallmark of aggressive cancers. Micronuclei frequently undergo irreversible collapse, exposing their enclosed chromatin to the cytosol. Micronuclear rupture catalyzes chromosomal rearrangements, epigenetic abnormalities, and inflammation, yet mechanisms safeguarding micronuclear integrity are poorly understood. In this study, we found that mitochondria-derived reactive oxygen species (ROS) disrupt micronuclei by promoting a noncanonical function of charged multivesicular body protein 7 (CHMP7), a scaffolding protein for the membrane repair complex known as endosomal sorting complex required for transport III (ESCRT-III). ROS retained CHMP7 in micronuclei while disrupting its interaction with other ESCRT-III components. ROS-induced cysteine oxidation stimulated CHMP7 oligomerization and binding to the nuclear membrane protein LEMD2, disrupting micronuclear envelopes. Furthermore, this ROS-CHMP7 pathological axis engendered chromosome shattering known to result from micronuclear rupture. It also mediated micronuclear disintegrity under hypoxic conditions, linking tumor hypoxia with downstream processes driving cancer progression.

Biochemical, genomic, and epigenomic profiling of isolated cancer cell lines' micronuclei.

Micronuclei are common byproducts of chromosomally unstable cancer cells during their division. Micronuclei play an important role in cancer metastasis as well as introducing chromosomal abnormalities into the primary nucleus in the subsequent cell cycle. Given their major role in tumor initiation and progression, methods to examine their properties are crucially needed. This chapter discusses approaches and methods to profile micronuclei's biochemical, genomic, and epigenomic properties following their physical isolation. Using either MPS1 inhibition or radiation to induce formation of micronuclei, this method introduces a versatile way to investigate biological events that occur in micronuclei as well as compare them to events in the primary nuclei from the same system.

Reversible histone glycation is associated with disease-related changes in chromatin architecture.

Cellular proteins continuously undergo non-enzymatic covalent modifications (NECMs) that accumulate under normal physiological conditions and are stimulated by changes in the cellular microenvironment. Glycation, the hallmark of diabetes, is a prevalent NECM associated with an array of pathologies. Histone proteins are particularly susceptible to NECMs due to their long half-lives and nucleophilic disordered tails that undergo extensive regulatory modifications; however, histone NECMs remain poorly understood. Here we perform a detailed analysis of histone glycation in vitro and in vivo and find it has global ramifications on histone enzymatic PTMs, the assembly and stability of nucleosomes, and chromatin architecture. Importantly, we identify a physiologic regulation mechanism, the enzyme DJ-1, which functions as a potent histone deglycase. Finally, we detect intense histone glycation and DJ-1 overexpression in breast cancer tumors. Collectively, our results suggest an additional mechanism for cellular metabolic damage through epigenetic perturbation, with implications in pathogenesis.