Probing barley mutants with a monoclonal antibody to a polypeptide involved in photosynthetic oxygen evolution

Abstract

A stable hybridoma line derived from the fusion of mouse myeloma cells with spleen cells from a mouse immunized with photosystem II vesicles has been isolated. The hybridoma cells secrete monoclonal antibodies to a 23 kD polypeptide present in isolated barley wild-type thylakoid membranes and known to be involved in water-splitting activity of photosystem II. No reaction between this monoclonal antibody to the 23 kD polypeptide and isolated internal etioplast membranes was detectable in immune-blot assays indicating that the synthesis of this polypeptide is stimulated by light. Immune-blots also revealed that the amount of 23 kD polypeptide in the photosystem II deficient mutantsviridis-zd 69 viridis-m 29 andviridis-115 is the same as in the wild-type. This indicates either that the oxygen evolving complex, of which the 23 kD polypeptide is a component, can be assembled independently of the photosystem II reaction centre, or that the 23 kD polypeptide, in the absence of the photosystem II reaction centre, is located in the thylakoid lumen and protected from protease attack. Furthermore it was shown that the cold sensitive barley mutanttigrina-o 34, contained the 23 kD polypeptide whether grown at 23°C or 30°C. The amount, on a chlorophyll basis, was reduced compared to that in the wild-type. Since the 23 kD polypeptide is present intigrina-o 34 plants grown at 23°C which lack chloroplast ribosomes, the 23 kD polypeptide must be coded for by a gene in the nucleus.