Relapse is the major cause of treatment-failure in adults with B-cell acute lymphoblastic leukemia (ALL) achieving complete remission after induction chemotherapy. Greater precision identifying persons likely to relapse is important. We did bio-informatics analyses of transcriptomic data to identify mRNA transcripts aberrantly-expressed in B-cell ALL. We selected 9 candidate genes for validation 7 of which proved significantly-associated with B-cell ALL. We next focused on function and clinical correlations of the cysteine and glycine-rich protein 2 (CSRP2). Quantitative real-time polymerase chain reaction (RT-qPCR) was used to examine gene transcript levels in bone marrow samples from 236 adults with B-cell ALL compared with samples from normals. CSRP2 was over-expressed in 228 out of 236 adults (97%) with newly-diagnosed B-cell ALL. A prognostic value was assessed in 168 subjects. In subjects with normal cytogenetics those with high CSRP2 transcript levels had a higher 5-year cumulative incidence of relapse (CIR) and worse relapse-free survival (RFS) compared with subjects with low transcript levels (56% [95% confidence interval, 53, 59%] vs. 19% [18, 20%]; P = 0.011 and 41% [17, 65%] vs. 80% [66–95%]; P = 0.007). In multivariate analyses a high CSRP2 transcript level was independently-associated with CIR (HR = 5.32 [1.64–17.28]; P = 0.005) and RFS (HR = 5.56 [1.87, 16.53]; P = 0.002). Functional analyses indicated CSRP2 promoted cell proliferation, cell-cycle progression, in vitro colony formation and cell migration ability. Abnormal CSRP2 expression was associated with resistance to chemotherapy; sensitivity was restored by down-regulating CSRP2 expression.

Eubacterium limosum ZL-II was described to convert secoisolariciresinol (SECO) to its demethylating product 4,4'-dihydroxyenterodiol (DHEND) under anoxic conditions. However, the reaction cascade remains unclear. Here, the O-demethylase being responsible for the conversion was identified and characterized. Nine genes encoding two methyltransferase-Is (MT-I), two corrinoid proteins (CP), two methyltransferase-IIs (MT-II), and three activating enzymes (AE) were screened, cloned, and expressed in Escherichia coli. Four of the nine predicted enzymes, including ELI_2003 (MT-I), ELI_2004 (CP), ELI_2005 (MT-II), and ELI_0370 (AE), were confirmed to constitute the O-demethylase in E. limosum ZL-II. The complete O-demethylase (combining the four components) reaction system was reconstructed in vitro. As expected, the demethylating products 3-demethyl-SECO and DHEND were both produced. During the reaction process, ELI_2003 (MT-I) initially catalyzed the transfer of methyl group from SECO to the corrinoid of ELI_2004 ([Co]-CP), yielding demethylating products and [CH-Co]-CP; then ELI_2005 (MT-II) mediated the transfer of methyl group from [CH-Co]-CP to tetrahydrofolate, forming methyltetrahydrofolate and [Co]-CP. Due to the low redox potential of [Co]/[Co], [Co]-CP was oxidized to [Co]-CP immediately in vitro, and ELI_0370 (AE) was responsible for catalyzing the reduction of [Co]-CP to its active form [Co]-CP. The active-site residues in ELI_2003, ELI_2005, and ELI_0370 were subsequently determined using molecular modeling combined with site-directed mutagenesis. To our knowledge, this is the first study on the identification and characterization of a four-component O-demethylase from E. limosum ZL-II, which will facilitate the development of method to artificial synthesis of related bioactive chemicals.