Characterization of two Arabidopsis thaliana myb-like proteins showing affinity to telomeric DNA sequence.
Telomere-binding proteins participate in forming a functional nucleoprotein structure at chromosome ends. Using a genomic approach, two Arabidopsis thaliana genes coding for candidate Myb-like telomere binding proteins were cloned and expressed in E. coli. Both proteins, termed AtTBP2 (accession Nos. T46051 (protein database) and GI:638639 (nucleotide database); 295 amino acids, 32 kDa, pI 9.53) and AtTBP3 (BAB08466, GI:9757879; 299 amino acids, 33 kDa, pI 9.88), contain a single Myb-like DNA-binding domain at the N-terminus, and a histone H1/H5-like DNA-binding domain in the middle of the protein sequence. Both proteins are expressed in various A. thaliana tissues. Using the two-hybrid system interaction between the proteins AtTBP2 and AtTBP3 and self interactions of each of the proteins were detected. Gel-retardation assays revealed that each of the two proteins is able to bind the G-rich strand and double-stranded DNA of plant telomeric sequence with an affinity proportional to a number of telomeric repeats. Substrates bearing a non-telomeric DNA sequence positioned between two telomeric repeats were bound with an efficiency depending on the length of interrupting sequence. The ability to bind variant telomere sequences decreased with sequence divergence from the A. thaliana telomeric DNA. None of the proteins alone or their mixture affects telomerase activity in vitro. Correspondingly, no interaction was observed between any of two proteins and the Arabidopsis telomerase reverse transcriptase catalytic subunit TERT (accession No. AF172097) using two-hybrid assay.
Schrumpfová
Petra
P
Department of Functional Genomics and Proteomics, Masaryk University Brno, Czech Republic.
Kuchar
Milan
M
Miková
Gabriela
G
Skrísovská
Lenka
L
Kubicárová
Tatiana
T
Fajkus
Jirí
J
D017360:Arabidopsis / Q000235:genetics; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D001667:Binding, Competitive; D003001:Cloning, Molecular; D024202:Electrophoretic Mobility Shift Assay; D015870:Gene Expression; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D020598:Proto-Oncogene Proteins c-myb / Q000235:genetics / Q000378:metabolism*; D019098:Telomerase / Q000378:metabolism; D016615:Telomere / Q000235:genetics / Q000378:metabolism; D034501:Telomere-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D029681:Arabidopsis Proteins; D010940:Plant Proteins; D020598:Proto-Oncogene Proteins c-myb; D019098:Telomerase; D034501:Telomere-Binding Proteins; C511988:ATR1 protein, Arabidopsis; C511989:CPC protein, Arabidopsis; C495844:TBP2 protein, Arabidopsis; C529793:At5g67580 protein, Arabidopsis