Jasna Puizina

Molecular analysis of telomere fusions in Arabidopsis: multiple pathways for chromosome end-joining.

End‐to‐end fusion of critically shortened telomeres in higher eucaryotes is presumed to be mediated by nonhomologous end‐joining (NHEJ). Here we describe two PCR‐based methods to monitor telomere length and examine the fate of dysfunctional telomeres in Arabidopsis lacking the catalytic subunit of telomerase (TERT) and the DNA repair proteins Ku70 and Mre11. Primer extension telomere repeat amplification relies on the presence of an intact G‐overhang, and thus measures functional telomere length. The minimum functional telomere length detected was 300–400 bp. PCR amplification and sequence analysis of chromosome fusion junctions revealed exonucleolytic digestion of dysfunctional ends prior to fusion. In ku70 tert mutants, there was a greater incidence of microhomology at the fusion junction than in tert mutants. In triple ku70 tert mre11 mutants, chromosome fusions were still detected, but microhomology at the junction was no longer favored. These data indicate that both Ku70 and Mre11 contribute to fusion of critically shortened telomeres in higher eucaryotes. Furthermore, Arabidopsis processes critically shortened telomeres as double‐strand breaks, using a variety of end‐joining pathways.

Arabidopsis SMG7 protein is required for exit from meiosis.

Meiosis consists of two nuclear divisions that are separated by a short interkinesis. Here we show that the SMG7 protein, which plays an evolutionarily conserved role in nonsense-mediated RNA decay (NMD) in animals and yeast, is essential for the progression from anaphase to telophase in the second meiotic division in Arabidopsis. Arabidopsis SMG7 is an essential gene, the disruption of which causes embryonic lethality. Plants carrying a hypomorphic smg7 mutation exhibit an elevated level of transcripts containing premature stop codons. This suggests that the role of SMG7 in NMD is conserved in plants. Furthermore, hypomorphic smg7 alleles render mutant plants sterile by causing an unusual cell-cycle arrest in anaphase II that is characterized by delayed chromosome decondensation and aberrant rearrangement of the meiotic spindle. The smg7 phenotype was mimicked by exposing meiocytes to the proteasome inhibitor MG115. Together, these data indicate that SMG7 counteracts cyclin-dependent kinase (CDK) activity at the end of meiosis, and reveal a novel link between SMG7 and regulation of the meiotic cell cycle.

Random amplified polymorphic DNA analysis, genome size, and genomic in situ hybridization of triploid viviparous onions

Triploid viviparous onions (Allium cepa L. var. viviparum Metzg. (ALEF.), auct.), (2n = 3x = 24), are known in some countries only as a rare relic crop, while in other parts of the world they are still traditionally or even commercially cultivated. Results indicating an identical random amplified polymorphic DNA (RAPD) banding pattern and the same DNA content (2C = 43.4 pg) establish the high genetic similarity and the unique origin of the Croatian clone Ljutika and the Indian clone Pran. In order to determine the parental Allium species of these natural triploid hybrids, genomic fluorescent in situ hybridization (GISH) was applied. Biotinylated genomic DNAs from six diploid Allium species (A. cepa L., A. fistulosum L., A. roylei Stearn, A. vavilovii M. Pop. et Vved., A. galanthum Kar. et Kir., A. oschaninii O. Fedtsch.) were used as probes in this study. While probes obtained from genomic DNA of A. cepa, A. vavilovii, and A. roylei hybridized to somatic chromosomes of Ljutika probes from A. fistulosum, A. galanthum, and A. oschaninii did not. The DNA probes of A. cepa and A. roylei each completely or predominantly labelled one genome (eight chromosomes). A few chromosomes, the markers of the triploid karyotype, were not completely labelled by any probe applied. Our GISH results indicate that triploid viviparous onions might possess a complex triparental genome organization.

Shell Characteristics of Land Snail Eobania vermiculata (Müller, 1774) (Helicidae) from Croatia

Abstract. n The land snail genus Eobania P. Hesse, 1913 in Croatia consists of four taxa: Eobania vermiculata vermiculata (O.F. Müller, 1774), E. v. pelagosana (Westerlund, 1894), E. v. figarole (Rensch, 1928) and E. v. kamenensis Berberović, 1963. Only E. v. vermiculata and E. v. pelagosana have been included in this study. The mean shell diameters (D) of the 13 samples ranged from 19.99 mm (sample from the island of Sušac) to 31.78 mm (Šibenik, seaside) and the mean shell height (H) from 17.31 mm (Sušac) to 25.66 mm (Island of Šolta). Shell morphology confirms that all island populations are smaller with examples of nanism on outside islands of Palagruža and Sušac. However, molecular analysis (16S rDNA and COI sequence data) showed no significant differences between the South Adriatic population from the mainland and islands, thus providing no molecular evidence for classification of a single E. vermiculata species into separate subspecies within the investigated area. The observed differences might possibly be attributed to ecological influences. The population with the largest average shell size was approximately 1.5x larger than the population with the smallest average size.

Fine-Scale Phylogeography of a Putative Secondary Contact Zone of the Land Snail Cornu aspersum (Gastropoda: Pulmonata: Helicidae) Along the Croatian Coast and Islands

Abstract: n Several morphological and molecular studies support the north African origin of the brown snail Cornu aspersum aspersum (Müller, 1774) and evidence suggests the presence of two morphologically and genetically different haplogroups in this area. However, all investigations were confined mainly to the western Mediterranean region, whereas the genetic structure and geographical distribution of eastern Mediterranean populations remained largely unknown. The present study attempted to elucidate the fine-scale phylogeography of C. a. aspersum along the Croatian coast and islands. Both Bayesian phylogenetic inference and network-based approaches showed a strong genetic splitting of Croatian 16S rRNA, COI and cyt b sequences into two well-defined groups consistent with previous eastern and western lineages. The eastern haplotypes were more frequent and distributed over a wider geographic area than western haplotypes. The genetic admixture of eastern and western gene pools observed in three Croatian populations would signify a new secondary contact zone in Croatia between both previously defined lineages. Our results suggest that: 1) the eastern lineage, which disappeared in most Mediterranean regions, persisted in the Adriatic area probably through several climatic cycles accumulating genetic differences and 2) two or, more likely, several independent post-glaciation colonization routes crossed at this contact zone, bringing two distinct C. a. aspersum haplogroups together.