Unusual evolutionary conservation of 5S rRNA pseudogenes inAspergillus nidulans: Similarity of the DNA sequence associated with the pseudogenes with the mouse immunoglobulin switch region

Summary AllAspergillus nidulans 5S rRNA pseudogenes known so far are the result of integration of an approx. 0.2-kbp-long DNA sequence into the 5S rRNA genes. This sequence, called block C, is present in at least five copies in theA. nidulans genome and seems to be associated either with 5S rRNA genes or pseudogenes. In contrast to the 78% sequence conservation of the C-block in pseudogenes, the truncated 5 halves of the pseudogenes are very highly conserved (96.9–100%). We postulate that the 5S rRNA pseudogenes are still a subject of concerted evolution. The C-block sequence shows similarity to the switch region of the mouse heavy chain immunoglobulin gene. A characteristic motif GGGTGAG is repeated several times in both sequences; the sequence conservation is 63%.     

Behavior of sperm nuclei in intact and bisected metaphase II mouse oocytes fertilized in the presence of colcemid

Our objective was to examine the developmental fate of sperm nuclei in oocytes fertilized under conditions of meiotic arrest. Therefore zona-free metaphase II oocytes and oocyte fragments (nucleate and anucleate) were fertilized in the presence of colcemid. In anucleate oocyte fragments, normal male pronuclei develop. In contrast, in intact oocytes and nucleate fragments sperm nuclei after initial decondensation undergo secondary condensation. This state is maintained as long as the oocytes are treated with colcemid. When the drug is removed 3 h after insemination, the meiotic spindle(s) is reconstructed, the second polar body(ies) is extruded, and a female pronucleus (or micronuclei) forms. At the same time the sperm nucleus decondenses again and transforms into a male pronucleus. In addition oocytes fertilized in the presence of colcemid could not be refertilized. These observations suggest that oocytes and oocyte fragments fertilized in the presence of colcemid undergo activation despite the failure of pronucleus formation. The inhibitory effect of colcemid on the formation of pronuclei is expressed only in the presence of oocyte chromosomes. We suggest that colcemid stabilizes factors responsible for chromosome condensation that are associated with oocyte chromosomes but not factors (whether the same or different) present in the cytoplasm.     

Transformation of sperm nuclei into male pronuclei in nucleate and anucleate fragments of parthenogenetic mouse eggs

Our objective was to examine the ability of nucleate and anucleate fragments of artificially activated mouse eggs to transform sperm nucleus into male pronucleus. To this end, zona-free oocytes in metaphase II were activated by ethanol and bisected into halves (one with the spindle, the other anucleate) either within 10 to 20 min (series A) or 3 or 5 hr later (series B). In series A, the fragments were inseminated 3,5, and 8 h after activation, and in series B. 3 and 5 h after activation. Both nucleate and anucleate fragments lose the capability of transforming sperm nucleus into fully formed pronucleus sometime between 3 and 5 h after activation. In 8 h old parthenogenetic fragments, the majority of sperm nuclei remain unchanged or begin decondensation but never reach the stage of an early pronucleus. In over 1/3 of anucleate fragments of this age group, sperm nuclei develop defectively: chromatin decondenses inside the persisting nuclear envelope. In other experimental groups, the incidence of these abnormal sperm nuclei varies between 0 and 10%. In general, the anuclcate fragments retain the capability to transform sperm nuclei (fully or partially) longer than their nuclear counterparts. This difference may be accounted for by a different level of substances required for pronuclcar growth (extrachromosomal constituents of the germinal vesicle and nuclear lamins): high and constant in the cytoplasm of anucleate egg halves and low and progressively decreasing in the nucleate halves because of their putative uptake by the female pronucleus. However, the cytoplasmic factors responsible for the initial stages of transformation (nuclear envelope breakdown, chromatin decondensation) become eventually inactivated both in the presence and in the absence of a female pronucleus.     

Autophagy as the cell survival in response to a microsporidian infection of the midgut epithelium of Isohypsibius granulifer granulifer (Eutardigrada: Hypsibiidae)

The midgut epithelial cells of many invertebrates may possess microorganisms which act as symbionts or pathogens (bacteria, microsporidia, viruses). During our previous studies on Isohypsibius granulifer granulifer Thulin, 1928 (Tardigrada, Eutardigrada), which examined alterations of the midgut epithelium during oogenesis, we found that some of the specimens were infected with microsporidia. All stages of pathogens occurred in the cytoplasm of the digestive cells in the midgut epithelium of I. g. granulifer that were infected with microsporidia: meronts, sporonts, sporoblasts, and spores. The cytoplasm of the digestive cells was rich in mitochondria, cisterns of rough endoplasmic reticulum (RER), and Golgi complexes. Autophagy in the digestive cells of the dorsal midgut was much more intensive in comparison with noninfected specimens. Membranes of phagophores surrounded the pathogens forming autophagosomes. These latter structures fused with lysosomes forming autolysosomes and residual bodies appeared. Neither glycogen granules nor droplets of varying electron density, which accumulated in digestive cells during vitellogenesis and choriogenesis, appeared in individuals with microsporidia. While the midgut epithelium in noninfected specimens takes part in vitellogenesis and choriogenesis, in infected specimens, midgut cells are involved in the process of autophagy as a survival strategy.     

Virulence properties of Campylobacter jejuni are enhanced by displaying a mycobacterial TlyA methylation pattern in its rRNA

Campylobacter jejuni is a bacterial pathogen that is generally acquired as a zoonotic infection from poultry and animals. Adhesion of C. jejuni to human colorectal epithelial cells is weakened after loss of its cj0588 gene. The Cj0588 protein belongs to the type I group of TlyA (TlyA^I) enzymes, which 2′‐O‐methylate nucleotide C1920 in 23S rRNA. Slightly longer TlyA^II versions of the methyltransferase are found in actinobacterial species including Mycobacterium tuberculosis, and methylate not only C1920 but also nucleotide C1409 in 16S rRNA. Loss of TlyA function attenuates virulence of both M. tuberculosis and C. jejuni. We show here that the traits impaired in C. jejuni null strains can be rescued by complementation not only with the original cj0588 (tlyA ^I) but also with a mycobacterial tlyA ^II gene. There are, however, significant differences in the recombinant phenotypes. While cj0588 restores motility, biofilm formation, adhesion to and invasion of human epithelial cells and stimulation of IL‐8 production in a C. jejuni null strain, several of these properties are further enhanced by the mycobacterial tlyA ^II gene, in some cases to twice the original wild‐type level. These findings strongly suggest that subtle changes in rRNA modification patterns can affect protein synthesis in a manner that has serious consequences for bacterial pathogenicity.     

New Strategies for Chemical Synthesis of Phosphorodithioates Derived from 2′- or 3′- or 5′-Thionucleosides

Abstract

Various phophorodithioates derived from thionucleosides were synthesis by the reaction anhydronucleosides with phosphorodithioic acids     

Tissue banking training courses: Polish experience

Personnel directly involved in the donation, procurement, testing, processing, preservation, storage and distribution of human tissues and cells should be appropriately qualified and provided with timely and relevant training according to EU directives. In the time of new tissue and cells regulations implementation such a training system existed in Poland only at a local level. The first training programme outlines for various groups of health professionals engaged in tissue banking practice was created in co-operation with the Institute for LifeLong Learning at University of Barcelona in 2006. This initial training courses were financially supported by EU Transition Facility Programme 2004. Then, starting from 2006, based on previous experience, system of advanced training courses was created. This training programme was financially supported by the National Programme for the Development of Transplantation Medicine 2006–2009—POLGRAFT financed by Polish Ministry of Health. During 2006 and 2007 first set of tissue banking initial training courses were provided according to TF 2004 project. Over 200 pathologists, forensic medicine specialists and other medical doctors responsible for donor screening and classification, medical directors of tissue establishments, technical staff; tissue graft users: orthopaedic surgeons, neurosurgeons, cardiosurgeons and ophthalmologists were trained. Between 2006 and 2009 there were organized 8 advanced tissue banking training courses according to POLGRAFT programme. There were organized both theoretical and practical courses on various aspects of tissue for over 350 persons. We present our experience in organisation of international and national tissue banking training courses.