Mutational analysis in podocin-associated hereditary nephrotic syndrome in Polish patients: founder effect in the Kashubian population

Hereditary nephrotic syndrome is caused by mutations in a number of different genes, the most common being NPHS2. The aim of the study was to identify the spectrum of NPHS2 mutations in Polish patients with the disease. A total of 141 children with steroid-resistant nephrotic syndrome (SRNS) were enrolled in the study. Mutational analysis included the entire coding sequence and intron boundaries of the NPHS2 gene. Restriction fragment length polymorphism (RFLP) and TaqMan genotyping assay were applied to detect selected NPHS2 sequence variants in 575 population-matched controls. Twenty patients (14 %) had homozygous or compound heterozygous NPHS2 mutations, the most frequent being c.1032delT found in 11 children and p.R138Q found in four patients. Carriers of the c.1032delT allele were exclusively found in the Pomeranian (Kashubian) region, suggesting a founder effect origin. The 14 % NPHS2 gene mutation detection rate is similar to that observed in other populations. The heterogeneity of mutations detected in the studied group confirms the requirement of genetic testing the entire NPHS2 coding sequence in Polish patients, with the exception of Kashubs, who should be initially screened for the c.1032delT deletion.     

Domain mapping of the Rad51 paralog protein complexes

The five human Rad51 paralogs are suggested to play an important role in the maintenance of genome stability through their function in DNA double-strand break repair. These proteins have been found to form two distinct complexes in vivo, Rad51B–Rad51C–Rad51D–Xrcc2 (BCDX2) and Rad51C–Xrcc3 (CX3). Based on the recent Pyrococcus furiosus Rad51 structure, we have used homology modeling to design deletion mutants of the Rad51 paralogs. The models of the human Rad51B, Rad51C, Xrcc3 and murine Rad51D (mRad51D) proteins reveal distinct N-terminal and C-terminal domains connected by a linker region. Using yeast two-hybrid and co-immunoprecipitation techniques, we have demonstrated that a fragment of Rad51B containing amino acid residues 1–75 interacts with the C-terminus and linker of Rad51C, residues 79–376, and this region of Rad51C also interacts with mRad51D and Xrcc3. We have also determined that the N-terminal domain of mRad51D, residues 4–77, binds to Xrcc2 while the C-terminal domain of mRad51D, residues 77–328, binds Rad51C. By this, we have identified the binding domains of the BCDX2 and CX3 complexes to further characterize the interaction of these proteins and propose a scheme for the three-dimensional architecture of the BCDX2 and CX3 paralog complexes.     

Carnitine: transport and physiological functions in the brain

Carnitine (4-N-trimethylammonium-3-hydroxybutyric acid), a compound necessary for a transfer of fatty acids for their oxidation within the cell, accumulates in brain although β-oxidation of fatty acids is very low in neurons. Carnitine accumulates to lower extent in the brain than in peripheral tissues and the mechanism of its transport through the blood–brain barrier is discussed, with the involvement of two transporters, OCTN2 and B^0,+ being presented. A limitation by the blood–brain barrier of carnitine supply for the brain and the mechanism of its transport to neural cells by a protein belonging to neurotransmitters' transporters superfamily is further discussed.

Due to the beneficial effects of administration of acetylcarnitine in case of patients with dementia, the role of this acylcarnitine is presented in the context of neuronal cell metabolism and the role of acetylcarnitine in the synthesis of acetylcholine. The roles of long-chain acyl derivatives of carnitine, in particular palmitoylcarnitine, responsible for interaction with the membranes, lipids acylation and specific interactions with proteins have been summarized. Stimulation of protein palmitoylation and a possibility of changing the acylation status of G proteins is described, as well as interaction of palmitoylcarnitine with protein kinase C. Diminished interaction of the isoform δ of this kinase with GAP-43 (B-50, neuromodulin), whose expression increases upon accumulation of either carnitine or palmitoylcarnitine points to a possible regulation of differentiation by these compounds and their role in neuroregeneration.     

Synthesis and induction of apoptosis in B cell chronic leukemia by diosgenyl 2-amino-2-deoxy-beta-D-glucopyranoside hydrochloride and its derivatives

2-Acetamido-2-deoxy-D-glucose hydrochloride (D-glucosamine hydrochloride) has been used for the preparation of 1,3,4,6-tetra-O-acetyl-2-deoxy-2-trifluoroacetamido-beta- (4) and 2-tetrachlorophthalimido-alpha,beta-D-glucopyranose (6), which have been transformed into the appropriate bromides and the chloride. Both bromo and chloro sugars were used as a glycosyl donors for the glycosylation of diosgenin [(25R)-spirost-5-en-3beta-ol]. These condensations were conducted under mild conditions, using silver triflate as a promoter, and gave diosgenyl glycosides 9 and 12. Each of them was converted into diosgenyl 2-amino-2-deoxy-beta-D-glucopyranoside hydrochloride (11) and N-acylamido derivatives. The structures of all new glycosides were established by 1H and 13C NMR spectroscopy. These diosgenyl glycosides are the first saponins containing the D-glucosamine residue that have been synthesized. These compounds show promising antitumor activities. The synthetic saponins increase the number of apoptotic B cells, in combination with cladribine (2-CdA), that are isolated from chronic lymphotic leukemia (B-CLL) patients.     

Heterochromatic banding patterns on chromosomes of twelve weevil species (Insecta,Coleoptera, Curculionoidea: Apionidae,Curculionidae)

The C-banding patterns of twelve weevil species are presented. The obtained results confirm the existence of two groups of species: with a small or large amount of heterochromatin in the karyotype. The first group comprises seven species (Apionidae: Holotrichapion pisi; Curculionidae: Phyllobius urticae, Ph. pyri, Ph. maculicornis, Tanymecus palliatus, Larinodontes turbinatus, Cionus tuberculosus). In weevils with a small amount of heterochromatin, tiny grains on the nucleus in interphase are visible, afterwards in mitotic and meiotic prophase appearing as dark dots. The absence of C-bands does not indicate a lack of heterochromatin but heterochromatic regions are sometimes so small that the condensation is not visible during the cell cycle. The second group comprises five species (Otiorhynchus niger, O. morio, Polydrusus corruscus, Barypeithes chevrolati, Nedyus quadrimaculatus) which possess much larger heteropicnotic parts of chromosomes visible during all nuclear divisions. The species examined have paracentromeric C-bands on autosomes and the sex chromosome X, except for Otiorhynchus niger, which also has an intercalary bands on one pair of autososomes. All the species examined differ in the size of segments of constitutive heterochromatin. The y heterochromosome is dot-like and wholly euchromatic in all the studied species.     

Day/night food consumption in mice is strain and age-dependent

Food consumption was measured during the day (lights on) and the night (lights off) and compared between one outbred and 9 inbred strains of mice (CBA/Kw, C3H, DBA2, KP, BALB/c, C57BL, B10.Amst, B10.BR, B10.BR Y-del) in age groups 30-60, 60-90, 90-120, and more than 120 days. Outbred mice and animals from B10 sublines ate significantly more during nocturnal darkness. Day and night food consumption was similar in KP animals. In mice from the remaining strains there was an apparent age-related shift from nocturnal towards diurnal eating habits.     

Distribution of polymorphic forms at the porcine GH locus in a population of day-10 pig embryos

The present study describes an analysis of genotype and allele distribution at the porcine GH locus among day-10 pig embryos. Embryos were collected post mortem from 6 crossbred (Danish Landrace x Yorkshire) sows inseminated with mixed Duroc semen and individually frozen for later analysis. After extraction, DNA was subjected to PCR amplification and restriction analysis with Msp I and Hae II enzymes. The genotype frequencies were: Msp I CD 0.17, DD 0.83; and Hae II AA 0.33, AB 0.58; and BB 0.09. The Msp I CC genotype was not found among analysed embryos. To our knowledge, this is the first report on the genotype and allele distribution at the GH locus among early pig embryos.     

Binding studies of eukaryotic initiation factor eIF4E with novel mRNA dinucleotide cap analogues

Studies on the interaction of the murine translation initiation factor 4E with two new-synthesized cap-analogues, modified at C2' of 7-methylguanosine, have been performed by means of the fluorescence titration method. No difference in the binding affinity for eIF4E was observed compared with the "anti reversed" cap analogues, possessing the analogous modifications at C3'. Potential significance of the novel caps as research tools for examination of the nuclear cap binding complex CBC80/20 has been discussed.