Effect of exogenous proline and water stress on protein metabolism in normal and pro mutant root tips

The proline requiring mutants (pro) identifiable on the basis of their collapsed endosperm morphology are seedling lethal monogenic recessive mutants. Their growth is resumed if L-proline is added to the medium (Gavazzi et al., Theor. Appl. Genet. 46:338-395, 1975). Free and protein amino acid composition established after feeding root tips with 14C glutamate and 14C ornithine show that proline biosynthesis is not blocked in pro mutants (Dierks Ventling C. and Tonelli C., Plant Physiol. 69:130-134, 1982).

We now present preliminary data on the electrophoretic analysis and protein metabolism of newly synthesized proteins in root tips under normal and mild water stress (-8 BARS) conditions and in the presence or absence of 1mM L-proline. Excised root tips of pro1-1 mutants and normal siblings in W22 inbred line were grown for 24 hours on liquid media and then incubated for 22 hours in (3H) L-amino acids mixture or (35S) L-methionine. The mitochondrial enriched fraction was isolated according to Nagahashi and Hiraike (Plant Physiol. 69:546-548, 1982). SDS PAGE electrophoresis and fluorography of the newly synthesized proteins of the mitochondrial and soluble fractions were carried out according to Bertani et al. (Z. Pflanzenphysiol. 103:37-43, 1981).

The results obtained can be summarized as follows:

1. The presence of proline in the medium induces a change in the polypeptide pattern of soluble and mitochondrial enriched fraction in both mutant and normal siblings.

2. Mutant root tips show a significantly higher uptake and protein incorporation of the labeled amino acids.

3. During water stress the uptake, protein incorporation and efficiency of incorporation are reduced in normal root tips while in mutants they appear unaffected.

4. A comparison of the polypeptide pattern of the mitochondrial and soluble fraction in mutant and normal siblings discloses only minor quantitative changes. No major changes in the polypeptide pattern appear thus associated to the mutational event leading to proline requirement.

Further experiments are underway.

Chiara Tonelli and Alcide Bertani

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