Cloning and characterization of a pentatricopeptide protein encoding gene (LOJ) that is specifically expressed in lateral organ junctions in Arabidopsis thaliana.
A line exhibiting expression of beta-glucuronidase (GUS) in the lateral organ junctions and shoot apical meristem (SAM) was identified from a population of T-DNA tagged lines carrying a promoter-less GUS gene. Southern hybridization confirmed the presence of a single T-DNA insertion in this line. The plant sequences flanking the T-DNA were cloned by TAIL PCR and sequenced. The insertion of T-DNA was found to be in the upstream region of a hypothetical gene (At2g39230). This gene, which we term as LOJ to indicate its specific expression in all lateral organ junctions encodes a predicted protein containing pentatricopeptide (PPR) motifs. This gene appears to belong to a group of TATA-less promoters and codes for a long ORF without any intron. The gene apparently codes for a protein of 97.65 kD with a mitochondrial target sequence at the N-terminal. Transcript analysis revealed that the expression of the gene is specifically restricted to the lateral organ junctions throughout the life of the plants. 5' RACE analysis revealed a 95 nucleotide long UTR region for this hypothetical gene. In silico analysis of the upstream region failed to identify a TATA box within -146 nucleotides. GUS expression analysis of the line 149 and the transgenic plants generated with constructs carrying the upstream sequences of this gene fused to uidA identified that the specificity of the expression of this gene resides within -569 to -152 bp region. The specific expression of LOJ at the base of lateral organ and shoot apical meristem (SAM) suggests an important role of LOJ in lateral organ development and boundary demarcation.
Prasad
Anand Mohan
AM
National Research Centre on Plant Biotechnology, Indian Agricultural Research Institute, New Delhi, India.
Sivanandan
C
C
Resminath
Radhamony
R
Thakare
Dhiraj R
DR
Bhat
S R
SR
Srinivasan
D017360:Arabidopsis / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics*; D015139:Blotting, Southern; D003001:Cloning, Molecular; D019295:Computational Biology / Q000379:methods; D004269:DNA, Bacterial / Q000235:genetics; D018744:DNA, Plant / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D020869:Gene Expression Profiling; D018506:Gene Expression Regulation, Plant; D005966:Glucuronidase / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D024363:Transcription Initiation Site
D029681:Arabidopsis Proteins; D004269:DNA, Bacterial; D018744:DNA, Plant; C502596:LOJ protein, Arabidopsis; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C036483:T-DNA; D005966:Glucuronidase