Diversity and distribution of reptiles in Romania

The reptile fauna of Romania comprises 23 species, out of which 12 species reach here the limit of their geographic range. We compiled and updated a national database of the reptile species occurrences from a variety of sources including our own field surveys, personal communication from specialists, museum collections and the scientific literature. The occurrence records were georeferenced and stored in a geodatabase for additional analysis of their spatial patterns. The spatial analysis revealed a biased sampling effort concentrated in various protected areas, and deficient in the vast agricultural areas of the southern part of Romania. The patterns of species richness showed a higher number of species in the warmer and drier regions, and a relatively low number of species in the rest of the country. Our database provides a starting point for further analyses, and represents a reliable tool for drafting conservation plans.     

Optimized expression and purification of myristoylated human neuronal calcium sensor 1 in E. coli

We have developed a protocol to produce large quantities of high purity myristoylated and non-myristoylated neuronal calcium sensor 1 (NCS-1) protein. NCS-1 is a member of the neuronal calcium sensor (NCS) family and plays an important role in modulating G-protein signaling and exocytosis pathways in cells. Many of these functions are calcium-dependent and require NCS-1 to be modified with an N-terminal myristoyl moiety. In our system, a C-terminally 6x His-tagged variant of NCS-1 was co-expressed with yeast N-myristoyltransferase (NMT) in ZYP-5052 auto-induction media supplemented with sodium myristate (100-200 microM). With optimized growth conditions and a high capacity metal affinity purification scheme, >50mg of homogenous myristoylated NCS-1 is obtained from 1L of culture in a single step. The properties of the C-terminally tagged NCS-1 variants are indistinguishable from those reported for untagged NCS-1. Using this system, we have also isolated and characterized mutant NCS-1 proteins that have attenuated (NCS-1 E120Q) and abrogated (NCS-1 DeltaEF) ability to bind calcium. The large quantities of NCS-1 proteins isolated from small culture volumes of auto-inducible media will provide the necessary reagents for further biochemical and structural characterization. The affinity tag at the C-terminus of the protein provides a suitable reagent for easily identifying binding partners of the various NCS-1 constructs. Additionally, this method could be used to produce other recombinant proteins of the NCS family, and may be extended to express and isolate myristoylated variants of other proteins.     

Arylphthalazines as potent,and orally bioavailable inhibitors of VEGFR-2

A series of arylphthalazine derivatives were synthesized and evaluated as antagonists of VEGF receptor II (VEGFR-2). IM-094482 57, which was prepared in two steps from commercially available starting materials, was found to be a potent inhibitor of VEGFR-2 in enzymatic, cellular and mitogenic assays (comparable activity to ZD-6474). Additionally, 57 inhibited the related receptor, VEGF receptor I (VEGFR-1), and showed excellent exposure when dosed orally to female CD-1 mice.     

The C9orf72-SMCR8-WDR41 complex is a GAP for small GTPases

ABSTRACT

Massive expansions of the hexanucleotide in C9orf72 are the primary genetic origins of familial amyotrophic lateral sclerosis (ALS) and frontal temporal dementia (FTD). Current studies have found that this repeat sequence participates in the disease process by producing neurotoxic substances and reducing the level of C9orf72 protein; however, the progress in the functional study of C9orf72 is slow. Recently, a stable complex, consisting of C9orf72, SMCR8, and WDR41, has been implicated in regulating membrane trafficking and macroautophagy. We reported the cryo-electron microscopy (cryo-EM) structure of the C9orf72-SMCR8-WDR41 complex (CSW complex), unveiling that the CSW complex is a dimer of heterotrimers. Intriguingly, in the heterotrimer of the C9orf72-SMCR8-WDR41, C9orf72 interacts with SMCR8 in a manner similar to the FLCN-FNIP2 complex. Nevertheless, WDR41 is connected to the DENN domain of SMCR8 through its N-terminal β-strand and C-terminal helix but does not directly interact with C9orf72. Notably, the C9orf72-SMCR8 complex was demonstrated to act as a GAP for RAB8A and RAB11A in vitro.     

RETRACTED ARTICLE: ShRNA-mediated gene silencing of Heat shock protein 70 inhibits human colon cancer cell growth in vitro and in vivo

Molecular and Cellular Biochemistry -     

A soluble form of the glycolipid-anchored receptor for urokinase-type plasminogen activator is secreted from peripheral blood leukocytes from patients with paroxysmal nocturnal hemoglobinuria.

The cellular urokinase-type plasminogen-activator (uPA) receptor (uPAR) is a glycolipid-anchored membrane protein thought to be involved in pericellular proteolysis during cell migration and tumor invasion. In the present study, we have identified and characterized two soluble forms of uPAR which have retained their ligand-binding capability. One variant was generated in vitro by treatment of intact normal cells with either a phosphatidylinositol-specific phospholipase C (PLC) or endoproteinase Asp-N. The other soluble uPAR variant was secreted in vivo from peripheral blood leukocytes affected by the stem-cell disorder paroxysmal nocturnal hemoglobinuria (PNH), and was found in the plasma from these PNH patients as well as in the conditioned medium from cultured PNH leukocytes. Under normal conditions, we find no evidence for any shedding or secretion of a soluble uPA-binding counterpart to human uPAR in plasma. Unlike normal leukocytes, the PNH-affected cells do not express uPAR on the cell surface, although they do contain apparently normal levels of uPAR-specific mRNA. The secreted uPAR derived from PNH cells has a mobility in SDS/PAGE that is slightly higher than that of uPAR solubilized by PtdIns-specific PLC or detergent, but resembles that of a truncated, recombinant uPAR variant, which has its C-terminus close to the proposed glycolipid-attachment site, suggesting that the secreted protein has been proteolytically processed for glycolipid attachment. The presence in plasma from PNH patients of such a secreted, hydrophilic form of uPAR lends support to the hypothesis that the lesion underlying the PNH disorder resides either in glycolipid biosynthesis or in the function of an as-yet-unidentified transamidating enzyme assumed to cleave and assemble the truncated uPAR with the preformed glycolipid moiety.     

Nosocomial Infection in Adult Admissions with Hematological Malignancies Originating from Different Lineages: A Prospective Observational Study

Background

Nosocomial infection (NI) causes prolonged hospital stays, increased healthcare costs, and higher mortality among patients with hematological malignancies (HM). However, few studies have compared the incidence of NI according to the HM lineage.

Objective

To compare the incidence of NI according to the type of HM lineage, and identify the risk factors for NI.

Methods

This prospective observational study monitored adult patients with HM admitted for >48 hours to the General Hospital of the People''s Liberation Army during 2010–2013. Attack rates and incidences of NI were compared, and multivariable logistic regression was used to control for confounding effects.

Results

This study included 6,613 admissions from 1,922 patients. During these admissions, 1,023 acquired 1,136 NI episodes, with an attack rate of 15.47% and incidence of 9.6‰ (95% CI: 9.1–10.2). Higher rates and densities of NIs were observed among myeloid neoplasm (MN) admissions, compared to lymphoid neoplasm (LN) admissions (28.42% vs. 11.00%, P<0.001 and 11.4% vs. 8.4‰, P<0.001). NI attack rates in acute myeloid leukemia (AML) and myelodysplastic/myeloproliferative neoplasm (MDS/MPN) were higher than those in MDS (30.69% vs. 20.19%, P<0.001; 38.89% vs. 20.19%, P = 0.003). Attack rates in T/NK-cell neoplasm and B-cell neoplasm were higher than those in Hodgkin lymphoma (15.04% vs. 3.65%; 10.94% vs. 3.65%, P<0.001). Multivariable regression analysis indicated prolonged hospitalization, presence of central venous catheterization, neutropenia, current stem cell transplant, infection on admission, and old age were independently associated with higher NI incidence. After adjusting for these factors, MN admissions still had a higher risk of infection (odds ratio 1.34, 95% CI: 1.13–1.59, P<0.001).

Conclusion

Different NI attack rates were observed for HM from different lineages, with MN lineages having a higher attack rate and incidence than LN lineages. Special attention should be paid to MN admissions, especially AML and MDS/MPN admissions, to control NI incidence.