Distinct roles of carbohydrate esterase family CE16 acetyl esterases and polymer-acting acetyl xylan esterases in xylan deacetylation.

Mass spectrometric analysis was used to compare the roles of two acetyl esterases (AE, carbohydrate esterase family CE16) and three acetyl xylan esterases (AXE, families CE1 and CE5) in deacetylation of natural substrates, neutral (linear) and 4-O-methyl glucuronic acid (MeGlcA) substituted xylooligosaccharides (XOS). AEs were similarly restricted in their action and apparently removed in most cases only one acetyl group from the non-reducing end of XOS, acting as exo-deacetylases. In contrast, AXEs completely deacetylated longer neutral XOS but had difficulties with the shorter ones. Complete deacetylation of neutral XOS was obtained after the combined action of AEs and AXEs. MeGlcA substituents partially restricted the action of both types of esterases and the remaining acidic XOS were mainly substituted with one MeGlcA and one acetyl group, supposedly on the same xylopyranosyl residue. These resisting structures were degraded to great extent only after inclusion of α-glucuronidase, which acted with the esterases in a synergistic manner. When used together with xylan backbone degrading endoxylanase and ß-xylosidase, both AE and AXE enhanced the hydrolysis of complex XOS equally.

Prevalence and geographic distribution of bovine viral diarrhoea (BVD) infection in Finland 1993-1997.

Bulk milk samples from every herd supplying milk to dairies in Finland were examined for the presence of antibodies to BVD virus (BVDV) annually during 1993-1997. The highest prevalence, 0.99% in 1994, declined to 0.37% in 1996; however, this favourable trend appeared to discontinue in 1997, where the prevalence remained at 0.41%. In 1993, sera of all individual animals from bulk milk antibody-positive herds were examined for the presence of these antibodies. Since 1994, only sera of animals from herds with a bulk milk absorbance reading greater than 0.250 in the EIA test were examined individually. Three geographic foci of BVDV antibody-positive dairy herds were resolved in 1994, one in the north-western, another in the eastern and a diffuse third in the southern part of Finland. A distinct limiting of the spread was apparent in 1997. Beef cattle were also studied during 1993-1997; in 1993 breeding units, in 1994 mainly beef suckler herds and in 1995-1997 serum samples of beef animals at slaughter were examined for the presence of antibodies to BVDV. The prevalence of seropositive herds in 1993 and 1994 was 30.2% and 3.2%, respectively, while the prevalence among slaughter animals ranged 0.8-1.6%. Seronegative animals in herds with > 50% of seropositive animals were examined for the presence of BVD-virus. A total of 40 dairy herds and two beef herds with viraemic (persistently infected, PI) animals was encountered during 1993-1997. A comprehensive control programme and a more specific, cooperatively funded eradication programme for dairy cattle were launched in 1994. These programmes most probably contributed to the decline in prevalence during 1994-1996.

Increased turnover of collagen in abdominal aortic aneurysms, demonstrated by measuring the concentration of the aminoterminal propeptide of type III procollagen in peripheral and aortal blood samples.

PURPOSE: The pathogenesis of abdominal aortic aneurysm (AAA) involves many factors; elastin degradation is believed to lead to initial dilation, whereas changes in the collagen structure predispose the aneurysm to rupture. The major collagens in the aortic wall are types I and III. We set out here to determine whether changes in serum propeptide of type III procollagen (PIIINP), a biologically relevant marker of type III collagen turnover, could be associated with the characteristics of AAA. METHODS: The aminoterminal PIIINP and the carboxyterminal propeptide of type I collagen were measured by radioimmunoassay in 87 patients with AAA and 90 control subjects with aortodistal arteriosclerosis. The samples were taken from the peripheral blood and from the abdominal aorta at the levels of the diaphragm and the common iliac artery. RESULTS: Mean PIIINP concentrations were higher in patients with AAA than in control subjects (3.47 micrograms/L vs 2.73 micrograms/L, p < 0.0001), correlating positively with aneurysm diameter in the former (r = 0.27, p = 0.04) and with the maximum thickness of the intraluminal thrombus (r = 0.39, p = 0.003). The gradient in PIIINP between the upper and lower end of the abdominal aorta was significant in the AAA group (-0.30 microgram/L, range -0.20 to -0.50 vs -0.10 micrograms/L, range -0.20 to 0.30, p = 0.002). CONCLUSIONS: These studies indicate that the turnover of type III collagen is increased in patients with AAA.

Location and alternative splicing of type XIII collagen RNA in the early human placenta.

BACKGROUND: Type XIII collagen is widely distributed in the fetus. It is characterized by complex alternative splicing of its primary transcripts in regions corresponding to nine exons of the gene. EXPERIMENTAL DESIGN: Localization of type XIII collagen mRNAs in early placentas was determined by in situ hybridization. Reverse-transcription-polymerase chain reactions were used to examine alternative splicing of nine exons in villous and decidual mRNAs. RESULTS: An intense in situ hybridization signal was observed in the fibroblastoid stromal cells of the placental villi. A moderate signal was found in the endothelial cells of developing capillaries and the cells of the cytotrophoblastic columns. Furthermore, mRNAs were detected in the large decidual cells of the decidual membrane and the stromal cells of the gestational endometrium, but not in the epithelial cells in the endometrial glands. Five combinations of exons 3B, 4A, 4B and 5, encoding half of the COL1 domain, were found. The combination lacking exons 3B-5 was the major variant in both villous and decidual mRNAs. Three combinations of exons 12 and 13 encoding the NC2 domain were found, the long variant containing either 12 or 13 sequences being the major variant in the villi while nearly equal amounts of long and short variants lacking both 12 and 13 sequences were observed in the decidua. Four variants of exons 29, 33 and 37, encoding parts of COL3 and NC4, were found as splicing out of exon 37 was not detected. The major variants in both mRNAs were two that lacked exon 29 and either lacked or contained exon 33 sequences. CONCLUSIONS: Type XIII collagen mRNAs were located in the placenta. Due to alternative splicing, the lengths of the COL1 and NC2 domains of this collagen vary from 57 to 104 and from 12 to 34 amino acids, respectively. The COL3 domain varies between 208 and 235 residues, whereas the NC4 is 18 residues.

Characterization of the spectrum of alternative splicing of alpha 1 (XIII) collagen transcripts in HT-1080 cells and calvarial tissue resulted in identification of two previously unidentified alternatively spliced sequences, one previously unidentified exon, and nine new mRNA variants.

Amplification of a COL1-encoding region of alpha 1 (XIII) collagen transcripts of HT-1080 cell RNA suggested that exon 3 of the alpha 1 (XIII) collagen gene, which was previously deduced to be of 35 base pairs (bp) may consist of a constitutive 8-bp exon and an alternatively spliced 27-bp exon, termed here exons 3A and 3B, respectively. Furthermore, a previously unidentified alternatively spliced Gly-Xaa-Yaa-encoding exon designated as 4B was found between the sequences encoded by exons 4, redesignated here as 4A and 5. Six of the 16 potential combinations of the four consecutive alternatively spliced exons 3B, 4A, 4B, and 5 were found to exist in mRNAs, and as a result, the length of the COL1 domain may vary between 57 and 104 amino acid residues. Most of the NC2 domain is encoded by the alternatively spliced exons 12 and 13. Where previous analysis of cDNAs indicated that mRNA variants exist that contain either exon 12 or 13 sequences, amplification studies indicated here that there are also variants that lack both exons 12 and 13 but none that contain both exons simultaneously. Thus, the predicted length of this domain is either 12, 31, or 34 residues. Analyses covering both the COL1 and NC2 domains demonstrate that at least 12 mRNA species exist through the alternations of exons 3B-5, 12, and 13.

Patterns of expression of the six alternatively spliced exons affecting the structures of the COL1 and NC2 domains of the alpha 1(XIII) collagen chain in human tissues and cell lines.

Reverse transcription-polymerase chain reactions and RNA protection experiments were used to examine alternative splicing of the six exons 3B-5, 12, and 13 affecting the COL1 and NC2 domains of type XIII collagen in seven human tissues and four cell lines. Distinct differences in the proportions of the variant mRNAs were found. With respect to the COL1 domain, all studied samples contained mRNAs corresponding to the shortest COL1 variants of 57 and 66 residues, with the former variant being prominent in most samples. Most of the samples also contained notable amounts of mRNAs that corresponded to the longest COL1 variants, mainly those of 104 and 95 residues. Particularly the extent of inclusion of exon 12 and 13 sequences, encoding most of the NC2 domain, varied according to the type of tissue or cell analyzed. Bone, cartilage, and colon adenocarcinoma samples contained little or none of the mRNAs corresponding to the long NC2 variants, whereas in fibroblast, lung, muscle, and osteosarcoma cells, those mRNAs were the major variants. The relative proportions of the various combinations of exons 3B-5, 12, and 13 were evaluated in four of the RNA samples. Interestingly, each of these samples appeared to contain only one to three major combinations of the six exons, representing about 40% to nearly 100% of all variants.