Submitted to: Rustbelt RNA Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 10/16/2008
Publication Date: 10/18/2008
Citation: Gaffney, B., Dinkins, R.D., Hunt, A.G. 2008. CPSF30 in Medicago. Rustbelt RNA Meeting. September 15, 2008, Deer Creek State Park and Resort, Mt. Sterling, OH. Poster #19.
Technical Abstract: CPSF30 is a component in 3’ RNA processing that has been well characterized in the model species Arabidopsis thaliana. In this study the gene for CPSF30 has been cloned out of Medicago truncatula and Medicago sativa. A large region within the gene exhibits significant conservation between A. thaliana and both medicago species. This area of the gene was cloned out and 5’ and 3’ RACE were used to get each full length gene. The Medicago genes were cloned into a protein expression vector. The vector was used to translate each gene into its corresponding protein and each protein was purified and dialyzed. CPSF30 in A. thaliana binds calmodulin and it was hypothesized that CPSF30 from both the Medicago species would also bind calmodulin based on the conservation of the amino acids between the model species and the Medicago species. This study shows that CPSF30 from both Medicago species does bind calmodulin. CPSF30 from A. thaliana also has been shown to possess three CCCH type zinc fingers, with the first implicated in RNA binding and the third in RNA cleavage. Amino acid alignments showed the zinc finger regions were conserved and it was hypothesized that CPSF30 from both Medicago species would bind RNA and cleave RNA. Electromobility shift assays were used to determine that RNA binding did occur. RNA nuclease activity in Medicago is still being tested. CPSF30 knockout studies in A. thaliana have yielded some consistent phenotypes that are still being characterized and explored. One such characteristic is an elevated resistance to oxidative stress. Currently CPSF30 knockouts are being produced in Medicago sativa that will enable further characterization of CPSF30 in an agriculturally relevant crop.