Hereditary cystatin C amyloid angiopathy: identification of the disease-causing mutation and specific diagnosis by polymerase chain reaction based analysis

Summary Hereditary cystatin C amyloid angiopathy (HCCAA) is a dominantly inherited disease characterized by amyloidosis, dementia and fatal cerebral hemorrhage of young adults. A method for rapid and simple diagnosis of HCCAA is described. It is based upon oligonucleotide-directed enzymatic amplification of a 275-bp genomic DNA segment containing exon 2 of the cystatin C gene from a blood sample, followed by digestion of the amplification product with AluI. Loss of an AluI recognition site in the amplified DNA segment from HCCAA patients results in a deviating band-pattern at agarose gel electrophoresis, compared with that obtained from normal subjects or unaffected HCCAA family members. In a population of 9 patients with manifest HCCAA, 14 patients with other causes of brain hemorrhage and 16 healthy individuals, the diagnostic procedure displayed a sensitivity and specificity for HCCAA of 100%. Amplified DNA segments from 4 HCCAA patients of four different families were analyzed by nucleotide sequencing; the HCCAA-causing mutation in all families was found to be a single TA substitution in the codon for amino acid residue 68 of cystatin C.     

A comparative study on mouthpart morphology of certain larvae of Chironomini (Diptera: Chironomidae) with reference to the larval feeding habits

Mouthparts of six species of chironomid larvae were compared on both an intra- and inter-specific level. The species were: Chironomus riparius, Cryptochironomus defectus gr., Endochironomus albipennis, Glyptotendipes pallens, Microtendipes pedellus and Parachironomus arcuatus . The mouthpart morphology in C. defectus gr. and P. arcuatus is considerably different from the four other species and changes little between instars. Within C. riparius, E. albipennis, G. pallens and M. pedellus there is a considerable difference in the mouthpart morphology between the first instar and the remaining instars. The morphology of the mouthparts of the first instar of C. riparius, E. albipennis, G. pallens and M. pedellus resembles in many ways that found in C. defectus gr. and P. arcuatus .     

Mutation in the structural gene for release factor 1 (RF-1) of Salmonella typhimurium inhibits cell division.

A temperature-sensitive mutant of Salmonella typhimurium LT2 was isolated. At the nonpermissive temperature cell division stopped and multinucleated filaments were formed. DNA, RNA, or protein synthesis was not affected until after about two generations. Different physiological conditions, such as anaerobiosis and different growth media, suppress the division deficiency at high temperatures. Certain mutations causing a reduced polypeptide chain elongation rate also suppress the division deficiency. The mutation is recessive and shown to be in the structural gene for release factor I (prfA). DNA sequencing of both the wild-type (prfA+) and mutant (prfA101) allele revealed a GC-to-AT transition in codon 168. Like other known prfA mutants, prfA101 can suppress amber mutations. The division defect in the prfA101 mutant strain could not be suppressed by overexpression of the ftsQAZ operon. Moreover, at the nonpermissive temperature the mutant shows a normal heat shock and SOS response and has a normal ppGpp level. We conclude that the prfA101-mediated defect in cell division is not directed through any of these metabolic pathways, which are all known to affect cell division. We speculate that the altered release factor I induces aberrant synthesis of an unidentified protein(s) involved in the elaborate process of septation.     

Structure and expression of the gene encoding cystatin D, a novel human cysteine proteinase inhibitor

A new member of the human cystatin multigene family has been cloned from a genomic library using a cystatin C cDNA probe. The complete nucleotide sequence of a 4.3-kilobase DNA segment, containing a complete gene with structure very similar to those of known Family 2 cystatin genes, was determined. The novel gene, called CST4, is composed of three exons and two introns. It contains the coding information for a protein of 142 amino acid residues, which has been tentatively called cystatin D. The deduced amino acid sequence includes a putative signal peptide and presents 51-55% identical residues with the sequences of either cystatin C or the secretory gland cystatins S, SN, or SA. The cystatin D sequence contains all regions of relevance for cysteine proteinase inhibitory activity and also the 4 cysteine residues that form disulfide bridges in the other members of cystatin Family 2. Northern blot analysis revealed that the cystatin D gene is expressed in parotid gland but not in seminal vesicle, prostate, epididymis, testis, ovary, placenta, thyroid, gastric corpus, small intestine, liver, or gall-bladder tissue. This tissue-restricted expression is in marked contrast with the wider distribution of all the other Family 2 cystatins, since cystatin C is expressed in all these tissues and the secretory gland cystatins are present in saliva, seminal plasma, and tears. Cystatin D, being the first described member of a third subfamily within the cystatin Family 2, thus appears to have a distinct function in the body in contrast to other cystatins.     

Gastric cytoprotection by intracisternal interleukin-1 beta in the rat

The effects of intracisternal (ic) injection of recombinant interleukin-1 beta (IL-1) on absolute ethanol-induced gastric necrotic lesions were studied in conscious rats. IL-1 given ic inhibited ethanol-induced gastric lesions. The cytoprotective effect was dose dependent (ED175 ng/rat), long lasting with a maximal action when given 1-3 h prior to ethanol, blocked by ic injection of a IL-1 receptor antagonist protein (IRAP), and by intraperitoneal injection of indomethacin. IL-1, injected ic, was detected in the peripheral blood. However, IL-1 serum levels were lower after IL-1 injection ic than after ip at a dose giving equal gastric protection. These data show that ic IL-1 induces long lasting gastric protection mediated by interaction with IL-1 receptors and prostaglandin pathways at central and/or peripheral sites that remain to be localized.     

The influence of 5-halo substituents on the thermal depyrimidination of the glycosidic bond in 2'-deoxyuridines

Following fusion, 2′-deoxynucleosides undergo thermolytic cleavage of the glycosidic bond to yield the corresponding base, furfuryl alcohol, and water. Purine deoxynucleosides are more readily subject to thermal cleavage of the base than are pyrimidine deoxynucleosides, such as 2′-deoxyuridine. However, when the latter deoxynucleoside is substituted by halogens in the 5-position, the glycosidic bond is weakened and loss of halouracil is competitive with ease of depurination. The reduction in strength of the glycosidic bond is linearly related to the corresponding meta Hammett substituent constant.