Transcript Slide 1
Work package 1 (WP1) Topic: Cyanobacterial Dormant Forms in an Aquatic environment Objective: Establishment of dormant stage/ Induction of akinetes in cyanobacteria Task Deliverable # month 1. Determine conditions for the induction of akinetes D4 9 (M9) 2. Physiological processes involved in the induction of the dormant stage D8 30 (M30) 3. Compounds and metabolites associated with the dormant stage D8 30 (M30) 4. Development of molecular tools: DNA libraries. Gene expression during akinete formation (avaK, hetR) D5 D6 18 (M18) 27 (M27) 5. Quantitative analyses of the expression of specific genes by QPCR D7 35 (M35) 6. Summary and Report (M36) # D4 # D8 Basic studies to reveal conditions and mechanisms involved in induction of akinetes formation, dormancy and desiccation: Nutrient Depletion (P), Light, Temp, O2 (hypoxia), K Storage of metabolites [carbohydrates, cyanophycin (N)] – enzymes involved, e.g cyanophycine synthase Mature akinetes Respiration and Photosynthetic apparatus (is there pigments production?) O2 evolution/consumption To collect adequate material for further genomic, proteomic and metabolomic studies. 1. Akinete formation K > HL > P 2. Storage metabolites in akinetes Cyanophycin the storage protein found in cyanobacteria is found in akinetes of A. ovalisporum and is enriched in aspartic and arginine amino acids. 3. Photosynthesis apparatus Young akinetes perform photosynthetic capabilities as found for vegetative cells grown in akinete-induced media. Mature akinetes maintain residual photosynthetic activity even though some components of the photosynthetic apparatus appear to remain intact. # D5 4. Molecular approach Two genomic libraries were cloned into pCC1Fos. The number of clones per culture: 13,056 fosmids 15,360 fosmids A total of 3481 sequencing reads were made from both library fosmids, using primers T7 and M13(-28) Comparison between Anabaena genome and Aphanizomenon contigs led to the selection of nine fosmids that may contain candidate genes loci. Gene Function Fosmid AvaK Akinete marker sbfos01-3p12, sbfos02-6e7 HetR protease with DNA binding activity sbfos02-9m21,sbfos02-7e10 DevR two-component system, regulatory protein sbfos01-3d6, sbfos02-8h10 HepA ABC transporter sbfos01-9d6 CphA Cyanophycin synthetase sbfos01-4l16 CphB Cyanophycinase sbfos01-4l16 Kdp operon K+ transporting ATPase sbfos01-3f13