טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentFridman Svetlana
SubjectIsolation and Characterization of a Cyanophage Carrying
Photosystem I Genes
DepartmentDepartment of Biology
Supervisors Professor Oded Beja
Professor Debbie Lindell
Full Thesis textFull thesis text - English Version


Abstract

Unicellular cyanobacteria of the genera Synechococcus and Prochlorococcus are important contributors to primary production in the open oceans. Over the past decade, various photosynthesis-related genes have been found in viruses that infect cyanobacteria (cyanophages). About 90% of cyanophage isolates carry the PSII psbA gene coding for the D1 protein, and it is thought that its expression increases phage fitness. In addition to the psbA gene, many cyanophages contain genes encoding the PSII D2 protein (psbD) or different genes of the photosynthetic electron transport chain.  More recently, cyanobacterial core photosystem I (PSI) genes were identified on environmental DNA thought to come from cyanophages. Cyanophages containing PSI genes were hypothesized to shift host metabolism toward a cyclic photosynthetic mode. However, prior to this study, no virus with PSI genes had been isolated. Using a screening scheme devised specifically to detect these phages, we isolated the phage, P-TIM68, from phage-concentrates collected from the Line Islands (central Pacific Ocean) with Prochlorococcus MIT9515 as host. The existence of the PSI gene-cluster in this phage was confirmed by whole genome sequencing. The 197 kb long genome revealed the presence of a PSI gene cassette containing 7 genes: the fused gene psaJF followed by psaC, A, B, K, E and D genes. This is the exact arrangement previously observed in the metagenomics data. Host range analysis determined that the phage is host specific. In addition, transmission electron microscopy and PCR with primers for the portal protein (g20) established that the phage is a T4-like myovirus. Characterization of the life cycle of P-TIM68 revealed a latent period of 10 hours (with 70%-75% infection at best) and phage yield reaching a plateau after 24 hours. Phage PSI and PSII genes were expressed with middle phage genes (e.g. DNA polymerase) and transcript levels reached their highest levels 6-8 hours after infection.  Phage PSI and PSII peptides were detected in host membranes by mass spectrometry.  In order to determine whether host photosynthesis is affected during P-TIM68 infection we measured different photosynthetic parameters in infected and uninfected cells. Our results show that total electron flow through PSI was higher in infected cells. Prior to this study, our knowledge of viral PSI genes came from metagenomic data only. Here, for the first time we isolated a phage containing both PSI and PSII genes and showed that these genes are expressed during infection.  By isolating the phage, we established a system that will be used to test the influence of phage photosynthesis genes on cyanobacterial photosynthesis.