Chlorophyll b is synthesized from chlorophyll a and is found in the light-harvesting complexes of prochlorophytes, green algae, and both nonvascular and vascular plants. We have used conserved motifs from the chlorophyll a oxygenase (CAO) gene from Chlamydomonas reinhardtii to isolate a homologue from Arabidopsis thaliana. This gene, AtCAO, is mutated in both leaky and null chlorina1 alleles, and DNA sequence changes cosegregate with the mutant phenotype. AtCAO mRNA levels are higher in three different mutants that have reduced levels of chlorophyll b, suggesting that plants that do not have sufficient chlorophyll b up-regulate AtCAO gene expression. Additionally, AtCAO mRNA levels decrease in plants that are grown under dimlight conditions. We have also found that the six major Lhcb proteins do not accumulate in the null ch1-3 allele.In nonvascular and vascular plants, green algae, and prochlorophytes, chlorophyll (Chl) b is one of the light harvesting pigments that function to bring photons to the reaction centers. Chl b binds and stabilizes many of the light-harvesting complex (LHC) proteins but is not associated with the reaction center or the core Chl-binding proteins of photosystem (PS)I and PSII. Chl b is not essential for growth, and many mutants that lack Chl b have been isolated; one of the best studied is the chlorina f2 mutant of barley. Paper chromatography was first used to demonstrate a loss of Chl b in this yellow-green mutant (1), and later a near complete reduction of Chl b was shown by using HPLC (2). Studies of LHCII biogenesis in chlorina f2 demonstrated that Lhcb1 gene transcription, translation, chloroplast import, and insertion into the thylakoid membrane all occurred without Chl b; however, the insertion was transient, and the Lhcb1 was rapidly degraded (3, 4). Etioplast membranes supplemented with Zn-pheophorbide b were able to integrate thermolysin-resistant Lhcb1 (5), providing biochemical evidence that stable insertion of Lhcb proteins requires Chl b. It is possible that the synthesis of Chl b could contribute to the regulation of the size of LHCII, and for this reason, we are interested in the regulation of Chl b synthesis in higher plants.More recent studies involving the use of monospecific LHC antibodies have shown that some LHC proteins are indeed stable in chlorina f2 despite the 100-fold reduction in Chl b (6-10). These include the LHCI proteins Lhca1, Lhca2, and Lhca3 and the LHC II protein Lhcb5. Lhcb2, Lhcb3, and Lhcb4 are partially stable, whereas Lhca4, Lhcb1, and Lhcb6 do not accumulate. Independent studies confirmed this work, also demonstrating the stability of Lhcb3 in chlorina f2 (11,12).The role of Lhcb1, the major LHCII protein that binds 40% of the total Chl (13) in different aspects of photosynthesis has been evaluated by studying chlorina f2. The mutant has an increased PSII͞PSI ratio to compensate for the loss of photons that would normally be harvested by Lhcb1 (14). Even so, a redox-controlled thylakoid kinase is not activated in chlorina f2 because t...

Entamoeba histolytica is the cause of amebic colitis and liver abscess. This parasite induces apoptosis in host cells and utilizes exposed ligands such as phosphatidylserine to ingest the apoptotic corpses and invade deeper into host tissue. The purpose of this work was to identify amebic proteins involved in the recognition and ingestion of dead cells. A member of the transmembrane kinase family, phagosome-associated TMK96 (PATMK), was identified in a proteomic screen for early phagosomal proteins. Anti-peptide affinity-purified antibody produced against PATMK demonstrated that it was a type I integral membrane protein that was expressed on the trophozoite surface, and that co-localized with human erythrocytes at the site of contact. The role of PATMK in erythrophagocytosis in vitro was demonstrated by: (i) incubation of ameba with anti-PATMK antibodies; (ii) PATMK mRNA knock-down using a novel shRNA expression system; and (iii) expression of a carboxy-truncation of PATMK (PATMKΔ932). Expression of the carboxy-truncation of PATMKΔ932 also caused a specific reduction in the ability of E. histolytica to establish infection in the intestinal model of amebiasis, however these amebae retained the ability to cause hepatic abscesses when directly injected in the liver. In conclusion, PATMK was identified as a member of the TMK family that participates in erythrophagocytosis and is uniquely required for intestinal infection.

Plants can change the size of their light harvesting complexes in response to growth at different light intensities. Although these changes are small compared to those observed in algae, their conservation in many plant species suggest they play an important role in photoacclimation. A polyclonal antibody to the C-terminus of the Arabidopsis thaliana chlorophyllide a oxygenase (CAO) protein was used to determine if CAO protein levels change under three conditions which perturb chlorophyll levels. These conditions were: (1) transfer to shaded light intensity; (2) limited chlorophyll synthesis, and (3) during photoinhibition. Transfer of wild-type plants from moderate to shaded light intensity resulted in a slight reduction in the Chl a/b ratio, and increases in both CAO and Lhcb1 mRNA levels as well as CAO protein levels. CAO protein levels were also measured in the cch1 mutant, a P642L missense mutation in the H subunit of Mg-chelatase. This mutant has reduced total Chl levels and an increased Chl a/b ratio when transferred to moderate light intensity. After transfer to moderate light intensity, CAO mRNA levels decreased in the cch1 mutant, and a concomitant decrease in CAO protein levels was also observed. Measurements of tetrapyrrole intermediates suggested that decreased Chl synthesis in the cch1 mutant was not a result of increased feedback inhibition at higher light intensity. When wild-type plants were exposed to photoinhibitory light intensity for 3 h, total Chl levels decreased and both CAO mRNA and CAO protein levels were also reduced. These results indicate that CAO protein levels correlate with CAO mRNA levels, and suggest that changes in Chl b levels in vascular plants, are regulated, in part, at the CAO mRNA level.