Arginine methylation in subunits of mammalian pre-mRNA cleavage factor I

Mammalian cleavage factor I (CF I_m) is composed of two polypeptides of 25 kDa and either a 59 or 68 kDa subunit (CF I_m25, CF I_m59, CF I_m68). It is part of the cleavage and polyadenylation complex responsible for processing the 3′ ends of messenger RNA precursors. To investigate post-translational modifications in factors of the 3′ processing complex, we systematically searched for enzymes that modify arginines by the addition of methyl groups. Protein arginine methyltransferases (PRMTs) are such enzymes that transfer methyl groups from S-adenosyl methionine to arginine residues within polypeptide chains resulting in mono- or dimethylated arginines. We found that CF I_m68 and the nuclear poly(A) binding protein 1 (PABPN1) were methylated by HeLa cell extracts in vitro. By fractionation of these extracts followed by mass spectral analysis, we could demonstrate that the catalytic subunit PRMT5, together with its cofactor WD45, could symmetrically dimethylate CF I_m68, whereas pICln, the third polypeptide of the complex, was stimulatory. As sites of methylation in CF I_m68 we could exclusively identify arginines in a GGRGRGRF or “GAR” motif that is conserved in vertebrates. Further in vitro assays revealed a second methyltransferase, PRMT1, which modifies CF I_m68 by asymmetric dimethylation of the GAR motif and also weakly methylates the C-termini of both CF I_m59 and CF I_m68. The results suggest that native—as compared with recombinant—protein substrates may contain additional determinants for methylation by specific PRMTs. A possible involvement of CF I_m methylation in the context of RNA export is discussed.     

Can current native tree seedling production and infrastructure meet an increasing forest restoration demand in Brazil?

Recent global commitments have placed forest and landscape restoration at the forefront of countries' efforts to recover ecosystem services, conserve biodiversity, and mitigate the effects of climate change. However, it needs to be asked if current native tree seedling supply meets an increase in demand for forest restoration? This study assessed the current configuration, distribution, and production capacity of forest nurseries producing native trees in Brazil. Brazil provides an interesting example of how global agreements aligned with national policies can lead to the proliferation of native seedling nurseries, and the challenges faced to restore species‐rich native forest ecosystems. We found that the nurseries in the Atlantic Forest region can still meet an increase in demand—both in terms of seedling quantity and diversity—because most of their production capacity is not currently used. However, not all Brazilian biomes have sufficient nurseries to meet restoration demands, thus there is a risk of using native species from a few biogeographical regions in a much spatially wider and ecologically diverse area. In addition, lack of seed supply and qualified labor can hamper the growth of the market. Barriers to seed supply may also lead to low levels of genetic variability and floristic representation in the populations and ecosystems to be restored. We conclude that restoration of high‐diversity forest ecosystems requires policies and supportive programs, with emphasis on private nurseries, to guarantee adequate supply of native tree seedlings and provide the necessary incentives to develop the emergent economy of forest restoration.     

Auricular Chromoblastomycosis: A Case Report and Review of Published Literature

Subcutaneous chromoblastomycosis is an infection commonly seen in tropical and subtropical climates, usually caused by trauma with vegetables and often affects the host’s lower limbs. We report a case of auricular chromoblastomycosis in a 67-year-old man and discuss the rarity of this clinical manifestation of chromoblastomycosis in the medical literature. In the present case, the etiologic agent was Fonsecaea pedrosoi, the most common agent found in Brazil.     

Requirement of UNC93B1 reveals a critical role for TLR7 in host resistance to primary infection with Trypanosoma cruzi

UNC93B1 associates with TLR3, 7, and 9, mediating their translocation from the endoplasmic reticulum to the endolysosome, thus allowing proper activation by microbial nucleic acids. We found that the triple-deficient 3d mice, which lack functional UNC93B1 as well as functional endosomal TLRs, are highly susceptible to infection with Trypanosoma cruzi. The enhanced parasitemia and mortality in 3d animals were associated with impaired proinflammatory response, including reduced levels of IL-12p40 and IFN-γ. Importantly, the phenotype of 3d mice was intermediary between MyD88(-/-) (highly susceptible) and TLR9(-/-) (moderately susceptible), indicating the involvement of an additional UN93B1-dependent TLR(s) on host resistance to T. cruzi. Hence, our experiments also revealed that TLR7 is a critical innate immune receptor involved in recognition of parasite RNA, induction of IL-12p40 by dendritic cells, and consequent IFN-γ by T lymphocytes. Furthermore, we show that upon T. cruzi infection, triple TLR3/7/9(-/-) mice had similar phenotype than 3d mice. These data imply that the nucleic acid-sensing TLRs are critical determinants of host resistance to primary infection with T. cruzi.     

RioK1, a new interactor of protein arginine methyltransferase 5 (PRMT5), competes with pICln for binding and modulates PRMT5 complex composition and substrate specificity

Protein arginine methylation plays a critical role in differential gene expression through modulating protein-protein and protein-DNA/RNA interactions. Although numerous proteins undergo arginine methylation, only limited information is available on how protein arginine methyltransferases (PRMTs) identify their substrates. The human PRMT5 complex consists of PRMT5, WD45/MEP50 (WD repeat domain 45/methylosome protein 50), and pICln and catalyzes the symmetrical arginine dimethylation of its substrate proteins. pICln recruits the spliceosomal Sm proteins to the PRMT5 complex for methylation, which allows their subsequent loading onto snRNA to form small nuclear ribonucleoproteins. To understand how the PRMT5 complex is regulated, we investigated its biochemical composition and identified RioK1 as a novel, stoichiometric component of the PRMT5 complex. We show that RioK1 and pICln bind to PRMT5 in a mutually exclusive fashion. This results in a PRMT5-WD45/MEP50 core structure that either associates with pICln or RioK1 in distinct complexes. Furthermore, we show that RioK1 functions in analogy to pICln as an adapter protein by recruiting the RNA-binding protein nucleolin to the PRMT5 complex for its symmetrical methylation. The exclusive interaction of PRMT5 with either pICln or RioK1 thus provides the first mechanistic insight into how a methyltransferase can distinguish between its substrate proteins.     

Discovery of a novel family of CDK inhibitors with the program LIDAEUS: structural basis for ligand-induced disordering of the activation loop

A family of 4-heteroaryl-2-amino-pyrimidine CDK2 inhibitor lead compounds was discovered with the new database-mining program LIDAEUS through in silico screening. Four compounds with IC(50) values ranging from 17 to 0.9 microM were selected for X-ray crystal analysis. Two distinct binding modes are observed, one of which resembles the hydrogen bonding pattern of bound ATP. In the second binding mode, the ligands trigger a conformational change in the activation T loop by inducing movement of Lys(33) and Asp(145) side chains. The family of molecules discovered provides an excellent starting point for the design and synthesis of tight binding inhibitors, which may lead to a new class of antiproliferative drugs.     

Semaphorin 3F and Neuropilin-2 Control the Migration of Human T-Cell Precursors

Neuropilins and semaphorins are known as modulators of axon guidance, angiogenesis, and organogenesis in the developing nervous system, but have been recently evidenced as also playing a role in the immune system. Here we describe the expression and role of semaphorin 3F (SEMA3F) and its receptor neuropilin-2 (NRP2) in human T cell precursors. NRP2 and SEMA3F are expressed in the human thymus, in both lymphoid and non-lymphoid compartments. SEMA3F have a repulsive effect on thymocyte migration and inhibited CXCL12- and sphingosine-1-phosphate (S1P)-induced thymocyte migration by inhibiting cytoskeleton reorganization prior to stimuli. Moreover, NRP2 and SEMA3F are expressed in human T-cell acute lymphoblastic leukemia/lymphoma primary cells. In these tumor cells, SEMA3F also blocks their migration induced by CXCL12 and S1P. Our data show that SEMA3F and NRP2 are further regulators of human thymocyte migration in physiological and pathological conditions.     

Renal Function in Hepatosplenic Schistosomiasis – An Assessment of Renal Tubular Disorders

Background

Renal involvement in Schistosoma mansoni infection is not well studied. The aim of this study is to investigate the occurrence of renal abnormalities in patients with hepatosplenic schistosomiasis (HSS), especially renal tubular disorders.

Methods

This is a cross-sectional study with 20 consecutive patients with HSS followed in a medical center in Maceió, Alagoas, Brazil. Urinary acidification and concentration tests were performed using calcium chloride (CaCl₂) after a 12-h period of water and food deprivation. The biomarker monocyte chemoattractant protein 1 (MCP-1) was quantified in urine. Fractional excretion of sodium (FE_Na+), transtubular potassium gradient (TTKG) and solute-free water reabsorption (TcH₂O) were calculated. The HSS group was compared to a group of 17 healthy volunteers.

Results

Patients'' mean age and gender were similar to controls. Urinary acidification deficit was found in 45% of HSS patients. Urinary osmolality was significantly lower in HSS patients (588±112 vs. 764±165 mOsm/kg, p = 0,001) after a 12-h period of water deprivation. TcH₂O was lower in HSS patients (0.72±0.5 vs. 1.1±0.3, p = 0.04). Urinary concentration deficit was found in 85% of HSS patients. The values of MCP-1 were higher in HSS group than in control group (122±134 vs. 40±28 pg/mg-Cr, p = 0.01) and positively correlated with the values of microalbuminuria and proteinuria.

Conclusions

HSS is associated with important kidney dysfunction. The main abnormalities found were urinary concentrating ability and incomplete distal acidification defect, demonstrating the occurrence of tubular dysfunction. There was also an increase in urinary MCP-1, which appears to be a more sensitive marker of renal damage than urinary albumin excretion rate.