The Ighmbp2 helicase structure reveals the molecular basis for disease-causing mutations in DMSA1

Mutations in immunoglobulin µ-binding protein 2 (Ighmbp2) cause distal spinal muscular atrophy type 1 (DSMA1), an autosomal recessive disease that is clinically characterized by distal limb weakness and respiratory distress. However, despite extensive studies, the mechanism of disease-causing mutations remains elusive. Here we report the crystal structures of the Ighmbp2 helicase core with and without bound RNA. The structures show that the overall fold of Ighmbp2 is very similar to that of Upf1, a key helicase involved in nonsense-mediated mRNA decay. Similar to Upf1, domains 1B and 1C of Ighmbp2 undergo large conformational changes in response to RNA binding, rotating 30° and 10°, respectively. The RNA binding and ATPase activities of Ighmbp2 are further enhanced by the R3H domain, located just downstream of the helicase core. Mapping of the pathogenic mutations of DSMA1 onto the helicase core structure provides a molecular basis for understanding the disease-causing consequences of Ighmbp2 mutations.     

A Putative Calcium-Permeable Cyclic Nucleotide-Gated Channel, CNGC18, Regulates Polarized Pollen Tube Growth

A tip-focused Ca^2＋ gradient is tightly coupled to polarized pollen tube growth, and tip-localized influxes of extracellular Ca^2＋ are required for this process. However the molecular identity and regulation of the potential Ca^2＋ channels remains elusive. The present study has implicated CNGC18 （cyclic nucleotide-gated channel 18） in polarized pollen tube growth, because its overexpression induced wider and shorter pollen tubes. Moreover, CNGC18 overexpression induced depolarization of pollen tube growth was suppressed by lower extracellular calcium （[Ca^2＋]ex）. CNGC18-yellow fluorescence protein （YFP） was preferentially localized to the apparent post-Golgi vesicles and the plasma membrane （PM） in the apex of pollen tubes. The PM localization was affected by tip-localized ROP1 signaling. Expression of wild type ROP1 or an active form of ROP1 enhanced CNGC18-YFP localization to the apical region of the PM, whereas expression of RopGAP1 （a ROP1 deactivator） blocked the PM localization. These results support a role for PM-Iocalized CNGC18 in the regulation of polarized pollen tube growth through its potential function in the modulation of calcium influxes.     

Chicken riboflavin-binding protein. cDNA sequence and homology with milk folate-binding protein

The Rd gene is expressed in the livers and oviducts of laying hens and codes for the riboflavin-binding protein (RfBP) of egg yolk and egg white. A lambda gt11 cDNA library derived from chicken oviduct poly(A)+ RNA was screened with polyclonal rabbit antiserum to chicken RfBP. Positive clones were isolated and rescreened with a mixed oligonucleotide probe corresponding to residues 20-25 of the mature protein. The largest cDNA clone (969 base pairs) was subcloned into plasmid pIBI21, and the nucleotide sequence was determined by the dideoxynucleotide method. This clone contained the entire coding region for RfBP. The published amino acid sequence of the mature protein was confirmed. In addition, the following 17-residue signal peptide was deduced: Met-Leu-Arg-Phe-Ala-Ile-Thr-Leu-Phe-Ala-Val-Ile-Thr-Ser-Ser-Thr-Cys. Unexpectedly, the nucleotide sequence codes for 2 adjacent arginine residues at the carboxyl terminus that are not observed in the mature protein. The amino acid sequence of RfBP is homologous with bovine milk folate-binding protein. Eight of the nine pairs of cysteines involved in disulfide bonds in RfBP are conserved in folate-binding protein, as are all of the tryptophan residues. Sequence identity between homologous regions of these two vitamin-binding proteins is more than 30%.     

Preliminary X-ray study of p-cresol methylhydroxylase (flavocytochrome c) from Pseudomonas putida N.C.I.B. 9869

Single crystals of p-cresol methylhydroxylase, a flavocytochrome c from Pseudomonas putida, have been prepared. The crystals are orthorhombic, space group P212121 with unit cell parameters; a = 140.3 A, b = 130.6 A and c = 74.1 A. They contain a single non-symmetric dimer per asymmetric unit and diffract to at least 2.5 A resolution.     

Anti-Inflammatory Effects of Acupuncture Stimulation via the Vagus Nerve

Although acupuncture therapy is widely used in traditional Asian medicine for the treatment of diverse internal organ disorders, its underlying biological mechanisms are largely unknown. Here, we investigated the functional involvement of acupuncture stimulation (AS) in the regulation of inflammatory responses. TNF-α production in mouse serum, which was induced by lipopolysaccharide (LPS) administration, was decreased by manual acupuncture (MAC) at the zusanli acupoint (stomach36, ST36). In the spleen, TNF-α mRNA and protein levels were also downregulated by MAC and were recovered by using a splenic neurectomy and a vagotomy. c-Fos, which was induced in the nucleus tractus solitarius (NTS) and dorsal motor nucleus of the vagus nerve (DMV) by LPS and electroacupuncture (EAC), was further increased by focal administration of the AMPA receptor blocker CNQX and the purinergic receptor antagonist PPADS. TNF-α levels in the spleen were decreased by CNQX and PPADS treatments, implying the involvement of inhibitory neuronal activity in the DVC. In unanesthetized animals, both MAC and EAC generated c-Fos induction in the DVC neurons. However, MAC, but not EAC, was effective in decreasing splenic TNF-α production. These results suggest that the therapeutic effects of acupuncture may be mediated through vagal modulation of inflammatory responses in internal organs.