Telomere length in paroxysmal nocturnal hemoglobinuria correlates with clone size

OBJECTIVE: To study if telomere length can be used as a surrogate marker for the mitotic history in normal and affected hematopoietic cells from patients with paroxysmal nocturnal hemoglobinuria (PNH). METHODS: The telomere length was measured by automated multicolor flow fluorescence in situ hybridization in glycosyl-phosphatidyl-inositol anchored protein (GPI)-negative and GPI-positive peripheral blood leukocytes. Eleven patients were studied, two with predominantly hemolytic PNH and nine with PNH associated with marrow failure. RESULTS: Telomere length in GPI-negative cells was significantly shorter than in GPI-positive cells of the same patient (p < 0.01, n = 11). The difference in telomere length (telomere length in GPI-positive minus telomere length in GPI-negative cells) correlated with the percentage of GPI-negative white blood cells. CONCLUSION: Our results support the hypothesis that telomere length is correlated to the replicative history of GPI-positive and GPI-negative cells and warrant further studies of telomere length in relation to disease progression in PNH.

Human intestinal maltase-glucoamylase: crystal structure of the N-terminal catalytic subunit and basis of inhibition and substrate specificity

Human maltase-glucoamylase (MGAM) is one of the two enzymes responsible for catalyzing the last glucose-releasing step in starch digestion. MGAM is anchored to the small-intestinal brush-border epithelial cells and contains two homologous glycosyl hydrolase family 31 catalytic subunits: an N-terminal subunit (NtMGAM) found near the membrane-bound end and a C-terminal luminal subunit (CtMGAM). In this study, we report the crystal structure of the human NtMGAM subunit in its apo form (to 2.0 A) and in complex with acarbose (to 1.9 A). Structural analysis of the NtMGAM-acarbose complex reveals that acarbose is bound to the NtMGAM active site primarily through side-chain interactions with its acarvosine unit, and almost no interactions are made with its glycone rings. These observations, along with results from kinetic studies, suggest that the NtMGAM active site contains two primary sugar subsites and that NtMGAM and CtMGAM differ in their substrate specificities despite their structural relationship. Additional sequence analysis of the CtMGAM subunit suggests several features that could explain the higher affinity of the CtMGAM subunit for longer maltose oligosaccharides. The results provide a structural basis for the complementary roles of these glycosyl hydrolase family 31 subunits in the bioprocessing of complex starch structures into glucose.

Role of two UDP-Glycosyltransferases from the L group of arabidopsis in resistance against pseudomonas syringae

The role of the salicylic acid (SA) glycosides SA 2-O-β-D-glucose (SAG), SA glucose ester (SGE) and the glycosyl transferases UGT74F1 and UGT74F2 in the establishment of basal resistance of Arabidopsis against Pseudomonas syringae pv tomato DC3000 (Pst) was investigated. Both mutants altered in the corresponding glycosyl transferases (ugt74f1 and ugt74f2) were affected in their basal resistance against Pst. The mutant ugt74f1 showed enhanced susceptibility, while ugt74f2 showed enhanced resistance against the same pathogen. Both mutants have to some extent, altered levels of SAG and SGE compared to wild type plants, however, in response to the infection, ugt74f2 accumulated higher levels of free SA until 24 hpi compared to wild type plants while ugt74f1 accumulated lower SA levels. These SA levels correlated well with reduced expression in PR1 and EDS1 in ugt74f1. In contrast, ugt74f2 has enhanced expression of Enhanced Disease Susceptibility 1 (EDS1) but a strong reduction in the expression of several jasmonate (JA)-dependent genes. Bacterial infection interfered with the expression of Fatty Acid Desaturase (FAD), Lipoxygenase2 (LOX2), carboxyl methyltransferase1 (BSMT1) and 9-cis-epoxycarotenoid dioxygenase (NCED3) genes in ugt74f1, thus promoting an antagonistic effect with SA-signalling and leading to enhanced bacterial growth. UGT74F2 might be a target for bacterial effectors since bacterial mutants affected in effector synthesis were impaired in inducing UGT74F2 expression. These results suggest that UGT74F2 negatively influences the accumulation of free SA, hence leading to an increased susceptibility due to reduced SA levels and increased expression of the JA and ABA markers LOX-2, FAD and NCED-3.