Mechanism of action of bovine testicular hyaluronidase. Mapping of the active site.

The reactions of purified, homogeneous bovine testicular hyaluronidase have been studied with radioactively labeled oligomers of hyalobiuronic acid, (GlcUA-GlcNAc)n, as substrates and acceptors. Transglycosylation occurs by transfer of a glycosyl residue with retention of configuration from a leaving group to an acceptor. On the basis of detailed examination of cleavage and transglycosylation patterns for the trimer; comparison of trimer, tetramer, and polymer as substrates; comparison of acceptors; equilibrium binding; and other data, it is proposed that the enzyme's active site consists of five subsites for hyalobiuronate residues. In the terminology of Schechter, I., and Berger, A. ((1966) Biochemistry 5, 3371), these are s2-s1-s' 2-s3, where the reducing terminus is to the right, and cleavage occurs between s1 and s' 1. It is proposed that subsite s'2 has a high affinity for a substrate residue, while s1 and s'1 have low substrate affinity, and s2 and s' 3 are intermediate in affinity. This proposal is seen to have mechanistic implications. The reactions of several substrates show similar bell-shaped pH dependences, with optima in the region of pH 5 to 5.5.     

Attraction of Culex pipiens to uropygial gland secretions does not explain feeding preference for American robins

Culex pipiens, the endemic mosquito vector of West Nile virus in eastern North America, is responsible for maintenance of the virus in avian reservoir hosts, the most important of which appears to be the American robin. One reason for the greater involvement of robins is believed to be the feeding preference of Cx. pipiens, however, the basis of this preference is not understood. We tested the hypothesis that the species‐specific chemical profile of avian uropygial gland secretions are used by Cx. pipiens as cues to locate birds and, therefore, may contribute to the observed feeding preferences. We used gas chromatography‐mass spectrometry to identify the semi‐volatile components of the uropygial gland secretions of American robins and two other common reservoir host species, the house sparrow and European starling. We found that the chemical composition of the robin secretions was different from those of the sparrows and starlings. Through behavioral choice trials conducted in a dual‐port olfactometer, we also found that Cx. pipiens did not prefer the secretions of robins over the other two species. Surprisingly, however, we found that Cx pipiens were more often attracted to live starlings over robins and to the secretions of starlings over those of robins.     

Molecular characterization and mapping of ALMT1, the aluminium-tolerance gene of bread wheat (Triticum aestivum L.).

The major aluminum (Al) tolerance gene in wheat ALMT1 confers. An Al-activated efflux of malate from root apices. We determined the genomic structure of the ALMT1 gene and found it consists of 6 exons interrupted by 5 introns. Sequencing a range of wheat genotypes identified 3 alleles for ALMT1, 1 of which was identical to the ALMT1 gene from an Aegilops tauschii accession. The ALMT1 gene was mapped to chromosome 4DL using 'Chinese Spring' deletion lines, and loss of ALMT1 coincided with the loss of both Al tolerance and Al-activated malate efflux. Aluminium tolerance in each of 5 different doubled-haploid populations was found to be conditioned by a single major gene. When ALMT1 was polymorphic between the parental lines, QTL and linkage analyses indicated that ALMT1 mapped to chromosome 4DL and cosegregated with Al tolerance. In 2 populations examined, Al tolerance also segregated with a greater capacity for Al-activated malate efflux. Aluminium tolerance was not associated with a particular coding allele for ALMT1, but was significantly correlated with the relative level of ALMT1 expression. These findings suggest that the Al tolerance in a diverse range of wheat genotypes is primarily conditioned by ALMT1.     

Cytochemistry of the skin of patients with mucopolysaccharidoses.

The distribution of complex carbohydrates has been investigated at the light and electron microscope levels in sweat glands of normal subjects and patients with Hurler's or Hunter's disease. Normal sweat glands examined with a battery of light microscopic histochemical methods revealed sulphated complex carbohydrate in secretory granules of the dark cells. These granules lacked affinity for dialysed iron (DI) at the light and electron microscope levels. The DI method demonstrated acid complex carbohydrates ultrastructurally on the surface of the intercellular canaliculi and central lumen in normal sweat glands. DI-reactive acidic material, presumably of mucopolysaccharide nature, surrounded and extended between collagen bundles in the stroma of normal skin, but was absent from the band which ensheathed the sweat gland and consisted of individual rather than bundled collagen fibrils. DI-reactive mucopolysaccharide lined and partially filled vacuoles of dark cells showing a laminar distribution in vacuoles of clear cells in sweat glands of a Hunter patient. The DI method also visualized mucopolysaccharide distributed throughout vacuoles in fibroblasts of this patient. DI-reactive acid material covered the luminal surface of the sweat gland, coated collagen bundles in the stroma and spared the periglandular collagenous sheath in skin from Hurler and Hunter patients as in that from normal controls. Acid phosphatase was localized ultrastructually in vacuoles and nearby cytoplasm and on plasmalemmae of clear cells, dark cells and myoepithelial cells of sweat glands from Hurler and Hunter patients. Vacuoles of dermal fibroblasts and Schwann cells in these specimens also exhibited strong acid phosphatase activity.     

Pathogenicity of Haemoproteus danilewskyi,Kruse, 1890, in blue jays (Cyanocitta cristata)

Although the impact of blood parasite infections on passerine birds is potentially great, little is known of their pathologic effects. We studied Haemoproteus danilewskyi in experimentally infected captive and naturally infected free-ranging blue jays (Cyanocitta cristata) to determine patterns of infection and examine the pathologic effects of the parasite on the host. Physiologic changes, such as elevated numbers of lymphocytes, heterophils, basophils, eosinophils, and monocytes and decreased packed cell volume in the peripheral blood were associated with the erythrocytic phase of experimental infections of captive juvenile jays. Sublethal pathologic changes associated with the pre-erythrocytic phase of infections were observed in the liver, lung, and spleen. Schizonts were observed in the pulmonary capillaries of a 1 yr old jay necropsied 31 days post-inoculation, but not in 20 juvenile jays necropsied 57 days post-inoculation. In free-ranging naturally infected jays plasma protein concentration increased with density of natural infections.     

Unique Antibody Responses to Malondialdehyde-Acetaldehyde (MAA)-Protein Adducts Predict Coronary Artery Disease

Malondialdehyde-acetaldehyde adducts (MAA) have been implicated in atherosclerosis. The purpose of this study was to investigate the role of MAA in atherosclerotic disease. Serum samples from controls (n = 82) and patients with; non-obstructive coronary artery disease (CAD), (n = 40), acute myocardial infarction (AMI) (n = 42), or coronary artery bypass graft (CABG) surgery due to obstructive multi-vessel CAD (n = 72), were collected and tested for antibody isotypes to MAA-modifed human serum albumin (MAA-HSA). CAD patients had elevated relative levels of IgG and IgA anti-MAA, compared to control patients (p<0.001). AMI patients had a significantly increased relative levels of circulating IgG anti-MAA-HSA antibodies as compared to stable angina (p<0.03) or CABG patients (p<0.003). CABG patients had significantly increased relative levels of circulating IgA anti-MAA-HSA antibodies as compared to non-obstructive CAD (p<0.001) and AMI patients (p<0.001). Additionally, MAA-modified proteins were detected in the tissue of human AMI lesions. In conclusion, the IgM, IgG and IgA anti-MAA-HSA antibody isotypes are differentially and significantly associated with non-obstructive CAD, AMI, or obstructive multi-vessel CAD and may serve as biomarkers of atherosclerotic disease.     

The molecular basis of potassium nutrition in plants

Over the last five years, the cloning and characterization of K⁺ transport genes corresponding to K⁺ channels (KAT1, AKT1, KST1, AKT2), associated subunits (KAB1) and a high-affinity transporter (HKT1) has opened up important new avenues for research on plant K⁺ nutrition. With the abundance of molecular data now available it seems timely to link this information with the wealth of data previously accumulated on the physiology of plant K⁺ acquisition. The ultimate goal of all this research is to gain a better understanding of K⁺ transport and nutrition in the intact plant. Thus it is important to begin to integrate the molecular research with results from biochemical and physiological research conducted at the cellular, root and whole plant levels. This article will focus on describing the features of the cloned K⁺ transporters and their possible roles in mediating high- and low-affinity K⁺ uptake from the soil, as well as how K⁺ acquisition may be regulated.Abbreviations NEM N-ethyl maleimide - PCMBS p-chloromercuribenzene sulphonic acid     

A human bone morphogenetic protein antagonist is down-regulated in renal cancer

We analyzed expression of candidate genes encoding cell surface or secreted proteins in normal kidney and kidney cancer. This screen identified a bone morphogenetic protein (BMP) antagonist, SOSTDC1 (sclerostin domain-containing-1) as down-regulated in kidney tumors. To confirm screening results, we probed cDNA dot blots with SOSTDC1. The SOSTDC1 message was decreased in 20/20 kidney tumors compared with normal kidney tissue. Immunohistochemistry confirmed significant decrease of SOSTDC1 protein in clear cell renal carcinomas relative to normal proximal renal tubule cells (p < 0.001). Expression of SOSTDC1 was not decreased in papillary and chromophobe kidney tumors. SOSTDC1 was abundantly expressed in podocytes, distal tubules, and transitional epithelia of the normal kidney. Transfection experiments demonstrated that SOSTDC1 is secreted and binds to neighboring cells and/or the extracellular matrix. SOSTDC1 suppresses both BMP-7-induced phosphorylation of R-Smads-1, -5, and -8 and Wnt-3a signaling. Restoration of SOSTDC1 in renal clear carcinoma cells profoundly suppresses proliferation. Collectively, these results demonstrate that SOSTDC1 is expressed in the human kidney and decreased in renal clear cell carcinoma. Because SOSTDC1 suppresses proliferation of renal carcinoma cells, restoration of SOSTDC1 signaling may represent a novel target in treatment of renal clear cell carcinoma.