The E262K mutation in Lamin A links nuclear proteostasis imbalance to laminopathy-associated premature aging.

Deleterious, mostly de novo, mutations in the lamin A (LMNA) gene cause spatio-functional nuclear abnormalities that result in several laminopathy-associated progeroid conditions. In this study, exome sequencing in a sixteen-year-old male with manifestations of premature aging led to the identification of a mutation, c.784G>A, in LMNA, resulting in a missense protein variant, p.Glu262Lys (E262K), that aggregates in nucleoplasm. While bioinformatic analyses reveal the instability and pathogenicity of LMNAE262K , local unfolding of the mutation-harboring helical region drives the structural collapse of LMNAE262K into aggregates. The E262K mutation also disrupts SUMOylation of lysine residues by preventing UBE2I binding to LMNAE262K , thereby reducing LMNAE262K degradation, aggregated LMNAE262K sequesters nuclear chaperones, proteasomal proteins, and DNA repair proteins. Consequently, aggregates of LMNAE262K disrupt nuclear proteostasis and DNA repair response. Thus, we report a structure-function association of mutant LMNAE262K with toxicity, which is consistent with the concept that loss of nuclear proteostasis causes early aging in laminopathies.

Methionine aminopeptidases with short sequence inserts within the catalytic domain are differentially inhibited: Structural and biochemical studies of three proteins from Vibrio spp.

Methionine aminopeptidases (MetAPs) have been recognized as drug targets and have been extensively studied for discovery of selective inhibitors. MetAPs are essential enzymes in all living cells. While most prokaryotes contain a single gene, some prokaryotes and all eukaryotes including human have redundancy. Due to the similarity in the active sites of the MetAP enzyme between the pathogens and human limited the success of discovering selective inhibitors. We recently have discovered that MetAPs with small inserts within the catalytic domain to have different susceptibilities against some inhibitors compared to those that do not have. Using this clue we used bioinformatic tools to identify new variants of MetAPs with inserts in pathogenic species. Two new isoforms were identified in Vibrio species with two and three inserts in addition to an isoform without any insert. Multiple sequence alignment suggested that inserts are conserved in several of the Vibrio species. Two of the three inserts are common between two and three insert isoforms. One of the inserts is identified to have "NNKNN" motif that is similar to well-characterized quorum sensing peptide, "NNWNN". Another insert is predicted to have a posttranslational modification site. Three Vibrio proteins were cloned, expressed, purified, enzyme kinetics established and inhibitor screening has been performed. Several of the pyridinylpyrimidine derivatives selectively inhibited MetAPs with inserts compared to those that do not have, including the human enzyme. Crystal structure and molecular modeling studies provide the molecular basis for selective inhibition.

Ectopic Expression of Rv0023 Mediates Isoniazid/Ethionamide Tolerance via Altering NADH/NAD+ Levels in Mycobacterium smegmatis.

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) accounts for nearly 1.2 million deaths per annum worldwide. Due to the emergence of multidrug-resistant (MDR) Mtb strains, TB, a curable and avertable disease, remains one of the leading causes of morbidity and mortality. Isoniazid (INH) is a first-line anti-TB drug while ethionamide (ETH) is used as a second-line anti-TB drug. INH and ETH resistance develop through a network of genes involved in various biosynthetic pathways. In this study, we identified Rv0023, an Mtb protein belonging to the xenobiotic response element (XRE) family of transcription regulators, which has a role in generating higher tolerance toward INH and ETH in Mycobacterium smegmatis (Msmeg). Overexpression of Rv0023 in Msmeg leads to the development of INH- and ETH-tolerant strains. The strains expressing Rv0023 have a higher ratio of NADH/NAD+, and this physiological event is known to play a crucial role in the development of INH/ETH co-resistance in Msmeg. Gene expression analysis of some target genes revealed reduction in the expression of the ndh gene, but no direct interaction was observed between Rv0023 and the ndh promoter region. Rv0023 is divergently expressed to Rv0022c (whiB5) and we observed a direct interaction between the recombinant Rv0023 protein with the upstream region of Rv0022c, confirmed using reporter constructs of Msmeg. However, we found no indication that this interaction might play a role in the development of INH/ETH drug tolerance.

A new approach for comprehensively describing heterogametic sex chromosomes.

Notwithstanding the rapid developments in sequencing techniques, Y and W sex chromosomes have still been mostly excluded from whole genome sequencing projects due to their high repetitive DNA content. Therefore, Y and W chromosomes are poorly described in most species despite their biological importance. Several methods were developed for identifying Y or W-linked sequences among unmapped scaffolds. However, it is not enough to discover functional regions from short unmapped scaffolds. Here, we provide a new and simple strategy based on k-mer comparison for comprehensive analysis of the W chromosome in Bombyx mori. Using this novel method, we effectively assembled de novo 1281 W-derived genome contigs (totaling 1.9 Mbp), and identified 156 W-linked transcript RNAs and 345 W-linked small RNAs. This method will help in the elucidation of mechanisms of sexual development and exploration of W chromosome biological functions, and provide insights into the evolution of sex chromosomes. Moreover, we showed this method can be employed in identifying heterogametic sex chromosomes (W and Y chromosomes) in many other species where genomic information is still scarce.