FIGURE 9.

CFM3 is associated with CAF1 and CAF2 in large particles in the chloroplast stroma. (A) Untreated (−RNase) and ribonuclease-A-treated (+ RNase) maize stromal extracts were sedimented through sucrose gradients under conditions in which particles greater than ∼70S pellet. An equal portion of each fraction was analyzed on immunoblots by probing with CFM3 or CAF2 antiserum. The CFM3 blot stained with Ponceau S is shown below to illustrate the position of Rubisco in the gradient (RbcL, large subunit of Rubisco). The modest effect of RNAse-treatment on the sedimentation of CFM3 particles is characteristic of group II intron ribonucleoprotein partices, which are resistant to RNase treatment (Till et al. 2001; Asakura and Barkan 2007; Watkins et al. 2007). CAF1 and RNC1 were shown previously to be recovered in analogous gradient fractions (Watkins et al. 2007). (B) CFM3 coimmunoprecipitates with CAF1 and CAF2. Chloroplast stroma was used for immunoprecipitations with the antibody indicated at the top. Antisera from two rabbits immunized with the CFM3 antigen were analyzed (lanes 1,2). Immunoblots of the immunoprecipitated proteins were probed with the antibodies indicated to the left.