An overview of chromatin modifications

The last 15 years have witnessed tremendous progress in elucidating the roles of chromatin modifications in transcription regulation, DNA repair, replication, recombination, and other genomic processes. In this issue of Biopolymers, a series of reviews will summarize recent advances in our understanding of chromatin modifying enzymes and explore unresolved questions with respect to their regulation and functions in gene expression and other nuclear processes. © 2012 Wiley Periodicals, Inc. Biopolymers 99: 95–97, 2013.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/94507/1/22158_ftp.pd

Discovery of cellular regulation by protein degradation

What follows is a story of some of the lab’s adventures mentioned above, including the inventions of new biochemical and genetic methods. This account stems, in part, from previous descriptions of the early history of the Ub field (31,32). Another antecedent is an interview I gave to Dr. Istvan Hargittai, a distinguished Hungarian chemist. It describes my life and science, including the early years in Moscow, the 1977 escape from the former Soviet Union, the essentially accidental hiring of me by MIT, and the work that ensued (33). The narrative below borrows from these sources, and mentions our more recent contributions as well

Parkinson's disease plasma biomarkers: An automated literature analysis followed by experimental validation

Diagnosis of Parkinson's disease (PD) is currently assessed by the clinical evaluation of extrapyramidal signs. The identification of specific biomarkers would be advisable, however most studies stop at the discovery phase, with no biomarkers reaching clinical exploitation. To this purpose, we developed an automated literature analysis procedure to retrieve all the background knowledge available in public databases. The bioinformatic platform allowed us to analyze more than 51,000 scientific papers dealing with PD, containing information on 4121 proteins. Out of these, we could track back 35 PD-related proteins as present in at least two published 2-DE maps of human plasma. Then, 9 different proteins (haptoglobin, transthyretin, apolipoprotein A-1, serum amyloid P component, apolipoprotein E, complement factor H, fibrinogen γ, thrombin, complement C3) split into 32 spots were identified as a potential diagnostic pattern. Eventually, we compared the collected literature data to experimental gels from 90 subjects (45 PD patients, 45 non-neurodegenerative control subjects) to experimentally verify their potential as plasma biomarkers of PD

Inactivation of the HR6B ubiquitin-conjugating DNA repair enzyme in mice causes male sterility associated with chromatin modification.

The ubiquitin-conjugating yeast enzyme RAD6 and its human homologs hHR6A and hHR6B are implicated in postreplication repair and damage-induced mutagenesis. The yeast protein is also required for sporulation and may modulate chromatin structure via histone ubiquitination. We report the phenotype of the first animal mutant in the ubiquitin pathway: inactivation of the hHR6B-homologous gene in mice causes male infertility. Derailment of spermatogenesis becomes overt during the postmeiotic condensation of chromatin in spermatids. These findings provide a parallel between yeast sporulation and mammalian spermatogenesis and strongly implicate hHR6-dependent ubiquitination in chromatin remodeling. Since heterozygous male mice and even knockout female mice are completely normal and fertile and thus able to transmit the defect, similar hHR6B mutations may cause male infertilit

The role of ubiquitination and hepatocyte growth factor-regulated tyrosine kinase substrate in the degradation of the adrenomedullin type I receptor

Calcitonin receptor-like receptor (CLR) and the receptor activity-modifying protein 2 (RAMP2) comprise a receptor for adrenomedullin (AM). Although it is known that AM induces internalization of CLR•RAMP2, little is known about the molecular mechanisms that regulate the trafficking of CLR•RAMP2. Using HEK and HMEC-1 cells, we observed that AM-induced activation of CLR•RAMP2 promoted ubiquitination of CLR. A mutant (CLRΔ9KR), lacking all intracellular lysine residues was functional and trafficked similar to the wild-type receptor, but was not ubiquitinated. Degradation of CLR•RAMP2 and CLRΔ9KR•RAMP2 was not dependent on the duration of AM stimulation or ubiquitination and occurred via a mechanism that was partially prevented by peptidase inhibitors. Degradation of CLR•RAMP2 was sensitive to overexpression of hepatocyte growth factor-regulated tyrosine kinase substrate (HRS), but not to HRS knockdown, whereas CLRΔ9KR•RAMP2 degradation was unaffected. Overexpression, but not knockdown of HRS, promoted hyperubiquitination of CLR under basal conditions. Thus, we propose a role for ubiquitin and HRS in the regulation of AM-induced degradation of CLR•RAMP2

Candidate Proteins, Metabolites and Transcripts in the Biomarkers for Spinal Muscular Atrophy (BforSMA) Clinical Study

Spinal Muscular Atrophy (SMA) is a neurodegenerative motor neuron disorder resulting from a homozygous mutation of the survival of motor neuron 1 (SMN1) gene. The gene product, SMN protein, functions in RNA biosynthesis in all tissues. In humans, a nearly identical gene, SMN2, rescues an otherwise lethal phenotype by producing a small amount of full-length SMN protein. SMN2 copy number inversely correlates with disease severity. Identifying other novel biomarkers could inform clinical trial design and identify novel therapeutic targets.To identify novel candidate biomarkers associated with disease severity in SMA using unbiased proteomic, metabolomic and transcriptomic approaches.A cross-sectional single evaluation was performed in 108 children with genetically confirmed SMA, aged 2-12 years, manifesting a broad range of disease severity and selected to distinguish factors associated with SMA type and present functional ability independent of age. Blood and urine specimens from these and 22 age-matched healthy controls were interrogated using proteomic, metabolomic and transcriptomic discovery platforms. Analyte associations were evaluated against a primary measure of disease severity, the Modified Hammersmith Functional Motor Scale (MHFMS) and to a number of secondary clinical measures.A total of 200 candidate biomarkers correlate with MHFMS scores: 97 plasma proteins, 59 plasma metabolites (9 amino acids, 10 free fatty acids, 12 lipids and 28 GC/MS metabolites) and 44 urine metabolites. No transcripts correlated with MHFMS.In this cross-sectional study, "BforSMA" (Biomarkers for SMA), candidate protein and metabolite markers were identified. No transcript biomarker candidates were identified. Additional mining of this rich dataset may yield important insights into relevant SMA-related pathophysiology and biological network associations. Additional prospective studies are needed to confirm these findings, demonstrate sensitivity to change with disease progression, and assess potential impact on clinical trial design.Clinicaltrials.gov NCT00756821

Biomarker candidates of neurodegeneration in Parkinson’s disease for the evaluation of disease-modifying therapeutics

Reliable biomarkers that can be used for early diagnosis and tracking disease progression are the cornerstone of the development of disease-modifying treatments for Parkinson’s disease (PD). The German Society of Experimental and Clinical Neurotherapeutics (GESENT) has convened a Working Group to review the current status of proposed biomarkers of neurodegeneration according to the following criteria and to develop a consensus statement on biomarker candidates for evaluation of disease-modifying therapeutics in PD. The criteria proposed are that the biomarker should be linked to fundamental features of PD neuropathology and mechanisms underlying neurodegeneration in PD, should be correlated to disease progression assessed by clinical rating scales, should monitor the actual disease status, should be pre-clinically validated, and confirmed by at least two independent studies conducted by qualified investigators with the results published in peer-reviewed journals. To date, available data have not yet revealed one reliable biomarker to detect early neurodegeneration in PD and to detect and monitor effects of drug candidates on the disease process, but some promising biomarker candidates, such as antibodies against neuromelanin, pathological forms of α-synuclein, DJ-1, and patterns of gene expression, metabolomic and protein profiling exist. Almost all of the biomarker candidates were not investigated in relation to effects of treatment, validated in experimental models of PD and confirmed in independent studies

The USP21 Short Variant (USP21SV) Lacking NES, Located Mostly in the Nucleus In Vivo, Activates Transcription by Deubiquitylating ubH2A In Vitro

USP21 is a deubiquitylase that catalyzes isopeptide bond hydrolysis between ubiquitin and histone H2A. Since ubiqutylated H2A (ubH2A) represses transcription, USP21 plays a role in transcriptional activation. On the other hand, the localization of USP21 suggests it has an additional function in the cytoplasm. Here, we identified a USP21 short variant (USP21SV) lacking a nuclear export signal (NES). Differential localization of USP21SV, more in the nucleus than the USP21 long variant (USP21LV), suggests they have redundant roles in the cell. Ectopic expression of both USP21 variants decreased ubH2A in the nucleus. Furthermore, both recombinant USP21 variants activate transcription by deubiquitylating ubH2A in vitro. These data suggest multiple roles for USP21 in the ubiquitylation-deubiquitylation network in the cell