Identification of BFN1, a Bifunctional Nuclease Induced during Leaf and Stem Senescence in Arabidopsis

Abstract

Nuclease I enzymes are responsible for the degradation of RNA and single-stranded DNA during several plant growth and developmental processes, including senescence. However, in the case of senescence the corresponding genes have not been reported. We describe the identification and characterization of BFN1 of Arabidopsis, and demonstrate that it is a senescence-associated nuclease I gene. BFN1 nuclease shows high similarity to the sequence of a barley nuclease induced during germination and a zinnia (Zinnia elegans) nuclease induced during xylogenesis. In transgenic plants overexpressing the BFN1 cDNA, a nuclease activity of about 38 kD was detected on both RNase and DNase activity gels. Levels of BFN1 mRNA were extremely low or undetectable in roots, leaves, and stems. In contrast, relatively high BFN1 mRNA levels were detected in flowers and during leaf and stem senescence. BFN1 nuclease activity was also induced during leaf and stem senescence. The strong response of the BFN1 gene to senescence indicated that it would be an excellent tool with which to study the mechanisms of senescence induction, as well as the role of the BFN1 enzyme in senescence using reverse genetic approaches in Arabidopsis.

RNS2, a conserved member of the RNase T2 family, is necessary for ribosomal RNA decay in plants

“…These mutants lack the main RNase activity found in Arabidopsis extracts. This RNase activity is present at high levels in all tissues of WT plants, and it is induced by Pi starvation and senescence, as described for the RNS2 transcript (7,20). Thus, we identified this band as RNS2 and confirmed that the T-DNA insertion produced a null mutation.…”

Cloning and Characterization of a cDNA Encoding a Putative Nuclease Related to Petal Senescence in Carnation (Dianthus caryophyllus L.) Flowers

“…-, gaps to align the amino acid sequence. GenBank Accession Nos: Arabidopsis thaliana BFN1, U90264 (Perez-Amador et al, 2000); Aspergillus oryzae nuclease S1, P24021 (Iwamatsu et al, 1991); carnation DcNUC1, AY986984 (this study); daffodil DAFSAG1, AF420010 (Hunter et al, 2002); daylily SA6, AF0832031 (Panavas et al, 1999); Penicillium citrinum nuclease P1, P24289 (Maekawa et al, 1991); Solanum tuberosum StEN1, AY676603; Zinnia ZEN1, AB003131 (Aoyagi et al, 1998).…”

“…The upstream primer was 5'-TCR GCS YTG TGY GTG TGG CCK GAY C-3' (nuclease multi F) and the downstream primer 5'-GGC YAG TCT NAC TCC WCC YTG NGC-3' (nuclease multi R). These primers were synthesized by referring to conserved nucleotide sequences among several nucleases of different origins (Perez-Amador et al, 2000). RT-PCR was performed according to the standard procedure with necessary optimization.…”

Flower senescence: some molecular aspects

“…Moreover, it has also been suggested that in Alstroemeria petals, PCD processes are initiated extremely early at a similar location on the petals to that observed for expression of the BFN1 promoter in tomato (wagstaff et al 2003). Cloning of the Arabidopsis BFN1 gene and sequencing of the corresponding polypeptide (protein) by Perez-amador et al (2000) have revealed the similarity of the BFN1 protein to DSA6 nuclease (involved in petal senescence; Panavas et al 1999) and ZEN1 nuclease (associated with PCD during tracheary element differentiation; Ito and Fukuda 2002). The regulation and expression pattern of BFN1 has been analyzed by cloning its 2.3 kb portion of the 5′ promoter sequence and then by detecting its ability to activate the GUS reporter gene construct.…”