Accumulation of plastid protein precursors under norflurazon-induced carotenoid deficiency and oxidative stress in barley

Young barley seedlings devoid of carotenoids were assayed for their ability to synthesize plastid heat and light stress proteins (HSP32, ELIP) under o xidative stress caused by norflurazon (NF). Dark-grown barley plantlets wer e grown from seeds imbibed in 10(-4) M NF, an inhibitor of carotenoid biosy nthesis, then, where indicated, subjected to heat stress (at 42 degreesC in the dark for 2 h). Thereafter the plantlets were either illuminated under a moderate high light flux (300 mu mol(.)m(-2.)s(-1)) or in low light (30 m u mol(.)m(-2.)s(-1)). Western blot analysis indicated that seedlings treate d with NF and heat shock contained in comparison with control seedlings two protein bands that could be detected with antibodies against small plastid HSPs: a mature protein HSP32 and an additional band of 36 kDa, while heat- shock-treated seedlings in the absence of NF contained merely HSP32. The am ount of 36-kDa protein increased with time of incubation under high light a nd after 24 h incubation its level exceeded that of HSP32. The data (immuno logical similarity, differences in the molecular masses, peptide mapping an d the disappearance of the 36-kDa band after removal of NF) suggested a pre cursor-product relationship between HSP32 and the 36-kDa protein. This 36-k Da precursor always associated with the membrane fraction while the mature protein resided partially in membranes, but predominantly in the soluble fr action of the cells. Similar effects of NF were shown on the accumulation o f the ELIP precursor. Thus, the accumulation of plastid protein precursors in NF-treated plantlets was shown both for the chloroplast HSP32, localized in stroma, and for FLIP localized in thylakoid membranes. A possible mecha nism for the accumulation of protein precursors is discussed. (C) 2001 Edit ions scientifiques et medicales Elsevier SAS.