Genetic and molecular analysis of T-DNA mutations in arabidopsis thaliana Dotaz Zobrazit nápovědu
Six different mutations with defects in immature seed development have been identified during screening of a T-DNA collection of Arabidopsis thaliana. The mutations were confirmed to be monogenic and recessive-lethal by genetic analysis. Mutant embryos were blocked in certain steps in the process necessary for embryo viability and development, and therefore they belong to the embryo-lethal class of mutants. The genetic and morphological studies of T-DNA mutations affecting embryo development are presented. The youngest embryos with a defect were observed at the globular stage in the VIII-64 mutation. Externally located cells, precursor of the protoderm, were characterised by abnormal cell division. VIII-41 mutation with a defect at the late globular stage was arrested at the globular-heart stage transition. VIII-111 mutation showed defect at heart stage of embryogenesis with atypical development of cotyledon primordia. The defect was associated with abnormal pattern of cell division constituting the precursor of the shoot apical meristem. In VIII-82 mutation defect in torpedo stage with asymmetric cotyledons was observed. Cotyledon stage of embryos and chlorophyll defect were observed in VIII-75 mutant. Abnormal suspensor consisting of two columns of cells was observed in 280-4-4 mutation. Newly identified embryo-lethals can serve as starting material for more detailed genetic and molecular studies.
- MeSH
- Agrobacterium tumefaciens MeSH
- Arabidopsis embryologie genetika MeSH
- buněčné dělení genetika MeSH
- DNA bakterií genetika MeSH
- fenotyp * MeSH
- inzerční mutageneze MeSH
- meristém cytologie MeSH
- mutace genetika MeSH
- semena rostlinná embryologie genetika MeSH
- zárodečné listy cytologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
- Názvy látek
- DNA bakterií MeSH
- T-DNA MeSH Prohlížeč
Analysis of plants bearing a T-DNA insertion is a potent tool of modern molecular biology, providing valuable information about the function and involvement of genes in metabolic pathways. A collection of 12 Arabidopsis thaliana lines with T-DNA insertions in the gene coding for the catalytic subunit of telomerase (AtTERT) and in adjacent regions was screened for telomerase activity [telomere repeat amplification protocol (TRAP) assay], telomere length (terminal restriction fragments), and AtTERT transcription (quantitative reverse transcription-PCR). Lines with the insertion located upstream of the start codon displayed unchanged telomere stability and telomerase activity, defining a putative minimal AtTERT promoter and the presence of a regulatory element linked to increased transcription in the line SALK_048471. Lines bearing a T-DNA insertion inside the protein-coding region showed telomere shortening and lack of telomerase activity. Transcription in most of these lines was unchanged upstream of the T-DNA insertion, while it was notably decreased downstream. The expression profile varied markedly in mutant lines harbouring insertions at the 5' end of AtTERT which showed increased transcription and abolished tissue specificity. Moreover, the line FLAG_385G01 (T-DNA insertion inside intron 1) revealed the presence of a highly abundant downstream transcript with normal splicing but without active telomerase. The role of regulatory elements found along the AtTERT gene is discussed in respect to natural telomerase expression and putative intron-mediated enhancement.
- MeSH
- Arabidopsis genetika MeSH
- DNA bakterií genetika MeSH
- genotyp MeSH
- inzerční mutageneze MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- proteiny huseníčku genetika MeSH
- regulace genové exprese u rostlin MeSH
- regulační oblasti nukleových kyselin genetika MeSH
- telomerasa genetika MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- DNA bakterií MeSH
- proteiny huseníčku MeSH
- T-DNA MeSH Prohlížeč
- telomerasa MeSH
- TERT protein, Arabidopsis MeSH Prohlížeč
The plant-specific receptor-like cytoplasmic kinases (RLCKs) form a large, poorly characterized family. Members of the RLCK VI_A class of dicots have a unique characteristic: their activity is regulated by Rho-of-plants (ROP) GTPases. The biological function of one of these kinases was investigated using a T-DNA insertion mutant and RNA interference. Loss of RLCK VI_A2 function resulted in restricted cell expansion and seedling growth. Although these phenotypes could be rescued by exogenous gibberellin, the mutant did not exhibit lower levels of active gibberellins nor decreased gibberellin sensitivity. Transcriptome analysis confirmed that gibberellin is not the direct target of the kinase; its absence rather affected the metabolism and signalling of other hormones such as auxin. It is hypothesized that gibberellins and the RLCK VI_A2 kinase act in parallel to regulate cell expansion and plant growth. Gene expression studies also indicated that the kinase might have an overlapping role with the transcription factor circuit (PIF4-BZR1-ARF6) controlling skotomorphogenesis-related hypocotyl/cotyledon elongation. Furthermore, the transcriptomic changes revealed that the loss of RLCK VI_A2 function alters cellular processes that are associated with cell membranes, take place at the cell periphery or in the apoplast, and are related to cellular transport and/or cell wall reorganisation.
- Klíčová slova
- Arabidopsis thaliana, cell expansion, gibberellins, hypocotyl growth, plant hormones, plant size, receptor-like cytoplasmic kinase, skotomorphogenesis, transcriptomic analysis,
- MeSH
- Arabidopsis účinky léků enzymologie genetika růst a vývoj MeSH
- DNA bakterií genetika metabolismus MeSH
- DNA vazebné proteiny genetika metabolismus MeSH
- geneticky modifikované rostliny MeSH
- gibereliny metabolismus farmakologie MeSH
- hypokotyl účinky léků enzymologie genetika růst a vývoj MeSH
- inzerční mutageneze MeSH
- kotyledon účinky léků enzymologie genetika růst a vývoj MeSH
- kyseliny indoloctové metabolismus farmakologie MeSH
- protein-serin-threoninkinasy genetika metabolismus MeSH
- proteiny huseníčku genetika metabolismus MeSH
- regulace genové exprese u rostlin * MeSH
- regulátory růstu rostlin farmakologie MeSH
- semenáček účinky léků enzymologie genetika růst a vývoj MeSH
- stanovení celkové genové exprese MeSH
- transkripční faktory bHLH genetika metabolismus MeSH
- transkripční faktory genetika metabolismus MeSH
- transkriptom MeSH
- vývojová regulace genové exprese MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- auxin response factor 6, Arabidopsis MeSH Prohlížeč
- BZR1 protein, Arabidopsis MeSH Prohlížeč
- DNA bakterií MeSH
- DNA vazebné proteiny MeSH
- gibereliny MeSH
- kyseliny indoloctové MeSH
- PIF4 protein, Arabidopsis MeSH Prohlížeč
- protein-serin-threoninkinasy MeSH
- proteiny huseníčku MeSH
- regulátory růstu rostlin MeSH
- T-DNA MeSH Prohlížeč
- transkripční faktory bHLH MeSH
- transkripční faktory MeSH
Sexual plant reproduction depends on the production and differentiation of functional gametes by the haploid gametophyte generation. Currently, we have a limited understanding of the regulatory mechanisms that have evolved to specify the gametophytic developmental programs. To unravel such mechanisms, it is necessary to identify transcription factors (TF) that are part of such haploid regulatory networks. Here we focus on bZIP TFs that have critical roles in plants, animals and other kingdoms. We report the functional characterization of Arabidopsis thaliana AtbZIP34 that is expressed in both gametophytic and surrounding sporophytic tissues during flower development. T-DNA insertion mutants in AtbZIP34 show pollen morphological defects that result in reduced pollen germination efficiency and slower pollen tube growth both in vitro and in vivo. Light and fluorescence microscopy revealed misshapen and misplaced nuclei with large lipid inclusions in the cytoplasm of atbzip34 pollen. Scanning and transmission electron microscopy revealed defects in exine shape and micropatterning and a reduced endomembrane system. Several lines of evidence, including the AtbZIP34 expression pattern and the phenotypic defects observed, suggest a complex role in male reproductive development that involves a sporophytic role in exine patterning, and a sporophytic and/or gametophytic mode of action of AtbZIP34 in several metabolic pathways, namely regulation of lipid metabolism and/or cellular transport.
- MeSH
- Arabidopsis genetika metabolismus MeSH
- buněčná stěna metabolismus ultrastruktura MeSH
- fluorescenční mikroskopie MeSH
- geneticky modifikované rostliny MeSH
- květy genetika růst a vývoj metabolismus MeSH
- metabolické sítě a dráhy genetika fyziologie MeSH
- mikroskopie elektronová rastrovací MeSH
- mutace MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- proteiny huseníčku genetika fyziologie MeSH
- pyl genetika metabolismus ultrastruktura MeSH
- pylová láčka genetika růst a vývoj metabolismus MeSH
- regulace genové exprese u rostlin MeSH
- stanovení celkové genové exprese MeSH
- testy genetické komplementace MeSH
- trans-aktivátory genetika fyziologie MeSH
- transmisní elektronová mikroskopie MeSH
- vývojová regulace genové exprese MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- bZIP34 protein, Arabidopsis MeSH Prohlížeč
- proteiny huseníčku MeSH
- trans-aktivátory MeSH