The current version of the androgen receptor (AR) gene mutations database is described. The total number of reported mutations has risen from 374 to 605, and the number of AR-interacting proteins described has increased from 23 to 70, both over the past 3 years. A 3D model of the AR ligand-binding domain (AR LBD) has been added to give a better understanding of gene structure-function relationships. In addition, silent mutations have now been reported in both androgen insensitivity syndrome (AIS) and prostate cancer (CaP) cases. The database also now incorporates information on the exon 1 CAG repeat expansion disease, spinobulbar muscular atrophy (SBMA), as well as CAG repeat length variations associated with risk for female breast, uterine endometrial, colorectal, and prostate cancer, as well as for male infertility. The possible implications of somatic mutations, as opposed to germline mutations, in the development of future locus-specific mutation databases (LSDBs) is discussed. The database is available on the Internet (http://www.mcgill.ca/androgendb/).

No abstract

The modification of glutamic acid residues to ␥-carboxyglutamic acid (Gla) is a post-translational modification catalyzed by the vitamin K-dependent enzyme ␥-glutamylcarboxylase. Despite ubiquitous expression of the ␥-carboxylation machinery in mammalian tissues, only 12 Gla-containing proteins have so far been identified in humans. Because bone tissue is the second most abundant source of Gla-containing proteins after the liver, we sought to identify Gla proteins secreted by bone marrow-derived mesenchymal stromal cells (MSCs). We used a proteomics approach to screen the secretome of MSCs with a combination of two-dimensional gel electrophoresis and tandem mass spectrometry. The most abundant Gla-containing protein secreted by MSCs was identified as periostin, a previously unrecognized ␥-carboxylated protein. In silico amino acid sequence analysis of periostin demonstrated the presence of four consensus ␥-carboxylase recognition sites embedded within fasciclin-like protein domains. The carboxylation of periostin was confirmed by immunoprecipitation and purification of the recombinant protein. Carboxylation of periostin could be inhibited by warfarin in MSCs, demonstrating its dependence on the presence of vitamin K. We were able to demonstrate localization of carboxylated periostin to bone nodules formed by MSCs in vitro, suggesting a role in extracellular matrix mineralization. Our data also show that another fasciclin I-like protein, ␤ig-h3, contains Gla. In conclusion, periostin is a member of a novel vitamin K-dependent ␥-carboxylated protein family characterized by the presence of fasciclin domains. Furthermore, carboxylated periostin is produced by bone-derived cells of mesenchymal lineage and is abundantly found in mineralized bone nodules in vitro.

Aims: The manner in which hydrogen sulfide (H 2 S) suppresses neuroinflammation is poorly understood. We investigated whether H 2 S polarized microglia to an anti-inflammatory (M2) phenotype by activating AMPactivated protein kinase (AMPK). Results: Three structurally unrelated H 2 S donors (5-(4-hydroxyphenyl)-3H-1,2-dithiocyclopentene-3-thione [ADT-OH], (p-methoxyphenyl) morpholino-phosphinodithioic acid [GYY4137], and sodium hydrosulfide [NaHS]) enhanced AMPK activation in BV2 microglial cells in the presence and absence of lipopolysaccharide (LPS). The overexpression of the H 2 S synthase cystathionine bsynthase (CBS) in BV2 cells enhanced endogenous H 2 S production and AMPK activation regardless of LPS stimulation. On LPS stimulation, overexpression of both ADT-OH and CBS promoted M2 polarization of BV2 cells, as evidenced by suppressed M1 and elevated M2 signature gene expression. The promoting effects of ADT-OH on M2 polarization were attenuated by an AMPK inhibitor or AMPK knockdown. Liver kinase B1 (LKB1) and calmodulin-dependent protein kinase kinase b (CaMKKb) are upstream kinases that activate AMPK. ADT-OH activated AMPK in Hela cells lacking LKB1. In contrast, both the CaMKKb inhibitor and siRNA abolished ADT-OH activation of AMPK in LPS-stimulated BV2 cells. Moreover, the CaMKKb inhibitor and siRNA blunted ADT-OH suppression on M1 gene expression and enhancement of M2 gene expression in LPS-stimulated BV2 cells. Moreover, ADT-OH promoted M2 polarization of primary microglia in an AMPK activation-and CaMKKb-dependent manner. Finally, in an LPS-induced in vivo neuroinflammation model, both ADT-OH and NaHS enhanced AMPK activation in the brain area where microglia were overactivated on LPS stimulation. Furthermore, ADT-OH suppressed M1 and promoted M2 gene expression in this in vivo model. Innovation and Conclusion: CaMKKb-dependent AMPK activation is an unrecognized mechanism underlying H 2 S suppression on neuroinflammation.