A Stimulatory Role for Cytokinin in the Arbuscular Mycorrhizal Symbiosis of Pea

The arbuscular mycorrhizal (AM) symbiosis between terrestrial plants and AM fungi is regulated by plant hormones. For most of these, a role has been clearly assigned in this mutualistic interaction; however, there are still contradictory reports for cytokinin (CK). Here, pea plants, the wild type (WT) cv. Sparkle and its mutant E151 (Pssym15), were inoculated with the AM fungus Rhizophagus irregularis. E151 has previously been characterized as possessing high CK levels in non-mycorrhizal (myc-) roots and exhibiting high number of fungal structures in mycorrhizal (myc+) roots. Myc- and myc+ plants were treated 7, 9, and 11 days after inoculation (DAI) with synthetic compounds known to alter CK status. WT plants were treated with a synthetic CK [6-benzylaminopurine (BAP)] or the CK degradation inhibitor INCYDE, whereas E151 plants were treated with the CK receptor antagonist PI-55. At 13 DAI, plant CK content was analyzed by mass spectrometry. The effects of the synthetic compounds on AM colonization were assessed at 28 (WT) or 35 (E151) DAI via a modified magnified intersections method. The only noticeable difference seen between myc- and myc+ plants in terms of CK content was in the levels of nucleotides (NTs). Whereas WT plants responded to fungi by lowering their NT levels, E151 plants did not. Since NTs are thought to be converted into active CK forms, this result suggests that active CKs were synthesized more effectively in WT than in E151. In general, myc+ and myc- WT plants responded similarly to INCYDE by lowering significantly their NT levels and increasing slightly their active CK levels; these responses were less obvious in BAP-treated WT plants. In contrast, the response of E151 plants to PI-55 depended on the plant mycorrhizal status. Whereas treated myc- plants exhibited high NT and low active CK levels, treated myc+ plants displayed low levels of both NTs and active CKs. Moreover, treated WT plants were more colonized than treated E151 plants. We concluded that CKs have a stimulatory role in AM colonization because increased active CK levels were paralleled with increased AM colonization while decreased CK levels corresponded to reduced AM colonization

Agro-morphological and molecular assessment of doubled haploid lines of pepper (Capsicum annuum L.)

Celem badań była ocena agromorfologiczna i molekularna pokoleń R3 ośmiu linii DH papryki (Capsicum annuum L.), otrzymanych w kulturach pylników mieszańców międzyodmianowych (ATZ1xPO)Fj oraz (ATZ1xCDT)Fr Opracowanie statystyczne wyników biometrycznych wykazało zróżnicowanie między badanymi formami, a także bardzo dużą jednorodność fenotypową w obrębie pojedynczej linii. Na szczególną uwagę zasługiwały linia AP10, plonująca na poziomie formy matecznej ATZ1, oraz linia AP15, która przewyższała pod tym względem obydwie formy rodzicielskie. Oceniane linie DH poddano analizie molekularnej za pomocą PCR-RAPD. Osiem starterów generowało produkty polimorficzne, co pozwoliło na rozróżnienie wszystkich ocenianych w doświadczeniu linii. Na podstawie otrzymanych elektroforogramów obliczono współczynniki dystansu genetycznego oraz opracowano dendrogramy ilustrujące zróżnicowanie genetyczne między formami rodzicielskimi i potomnymi liniami DH. Wyróżnione pod względem cech użytkowych linie AP10 i AP15 stanowiły wspólną grupę o najmniejszym współczynniku dystansu genetycznego.The aim of the research was the agro-morphological and molecular analysis of eight DH-R3 lines of pepper (Capsicum annuum L.). The following lines of C. annuum were used in the research - three parental forms: ATZ1, PO, CDT, and R3 generation of DH lines obtained in anther culture of two hybrids (ATZ1xPO)F1: lines AP1, AP2, AP10, AP15, AP32, AP40 and (ATZ1xCDT)F1: lines AC5, AC7. The seeds of R1 androgenic plants were used to obtain two further generations of regenerants. The assessment of the homogeneity of the DH-R3 lines was performed with the use of analysis of variations of the important biometrical traits of plants and fruits. Analysis of variance proved homogeneity of all the examined DH populations and Scheffe's test confirmed polymorphism between the lines of a common origin. The following lines deserved special attention: AP10 line - yielding at the level of the maternal form, and AP15 line, which surpassed both parental forms regarding yield level. Additionally, fruits of these two lines were characterized by thick pericarp and high content of dry matter. As biometrical characteristic of the agronomical traits do not always allow a clear identification of androgenic lines, a RAPD analysis was also performed. Genomic DNA of the tested lines was isolated from young leaves of healthy, adult plants. PCR was conducted according to Ilbi. 13 starters were used, which generated between 5 and 11 products of the size varied between 250 and 2350 base pairs. Reactions conducted with the use of the following starters: A01, A07, A10, A11, B10, E19, F05 and F10 generated between one and three polymorphic stripes, which allowed to distinguish and assess the similarity of all the examined lines. The coefficient of genetic distance for the tested lines was calculated using Nei and Li formula in Treecon. Dendrograms illustrating genetic diversity between the lines revealed similar genetic profile of both tested DH lines in the AC line group that displayed resemblance to the maternal ATZ1 line. In the AP group, AP32 and AP40 lines had a similar genetic profile to the maternal ATZ1 form, while AP1 and AP2 lines resembled the paternal form, PO line. AP10 and AP15 lines, previously distinguished with regards for the biometrical traits, comprised a common group of the lowest coefficient of genetic distance

Restriction endonucleases that cleave RNA DNA heteroduplexes bind dsDNA in A like conformation

Restriction endonucleases naturally target DNA duplexes. Systematic screening has identified a small minority of these enzymes that can also cleave RNA DNA heteroduplexes and that may therefore be useful as tools for RNA biochemistry. We have chosen AvaII G amp; 8595;GWCC, where W stands for A or T as a representative of this group of restriction endonucleases for detailed characterization. Here, we report crystal structures of AvaII alone, in specific complex with partially cleaved dsDNA, and in scanning complex with an RNA DNA hybrid. The specific complex reveals a novel form of semi specific dsDNA readout by a hexa coordinated metal cation, most likely Ca2 or Mg2 . Substitutions of residues anchoring this non catalytic metal ion severely impair DNA binding and cleavage. The dsDNA in the AvaII complex is in the A like form. This creates space for 2 amp; 8242; OH groups to be accommodated without intra nucleic acid steric conflicts. PD D E XK restriction endonucleases of known structure that bind their dsDNA targets in the A like form cluster into structurally similar groups. Most such enzymes, including some not previously studied in this respect, cleave RNA DNA heteroduplexes. We conclude that A form dsDNA binding is a good predictor for RNA DNA cleavage activit