Cloning Of Chloroplast Division Related Genes MinD, MinE And Expression Analysis Of MinD, MinE, FtsZ In Chlamydomonas Reinhardtii

Plastids are essential organelles of plant cells and it is believed that plastids have arisen from an endosymbiotic event between a protoeukaryote and a photosynthetic prokaryote. Plastids are not created de novo, but arise by a progress termed binary fission from pre-existing plastids. Plastids fission may share similarities with bacterial cell division because their prokaryotic origin. In bacteria cell division, FtsZ protein assembles into a ring structure (Z ring) at the mid-cell division site which possibly served to constrict the membrane. MinD, MinC and MinE proteins act in concert to control the assembly of FtsZ ring only at the cell center. FtsZ, MinD and MinE are prokaryote-derived key plastid division components identified in past years. These proteins act similar to their bacterial counterpart in plastid division. Different from a single ftsZ gene in bacteria, there are two distinct families of FtsZ in plants termed FtsZ1 and FtsZ2 respectively.The Chlamydomonas reinhardtii MinD gene and MinE gene assembled with ESTs were isolated by RT-PCR and designated CrMinD and CrMinE respectively. The full length of CrMinD cDNA is 1683bp containing an open reading frame(ORF) of 1056bp encoding a protein of 351 amino acids. Its N-terminal 66 amino acids are putative transitpeptide. The full length of CrMinE cDNA is 1627bp containing an ORF of 1047bp encoding a protein of 349 amino acids. Its N-terminal 52 amino acids are predicted to be the transitpeptide. Expressions of CrMinD and CrMinE in E.coli show that CrMinD and CrMinE still function in E. coli.The expression of CrFtsZ1, CrFtsZ2, CrMinD and CrMinE in transcript level in synchronous cell cycles of Chlamydomonas reihardtii were analyzed by semi-quantitative RT-PCR. The results showed that the level of these mRNAs fluctuated dramatically and share similar expression pattern in light/dark cycles, peaking at the light-dark transition point, that is just before plastid division. This expression pattern corresponds with the functions these proteins have in plastid division if the protein level follows the mRNA level. The mRNAs abundance rhythm persisted at least three cycles in continuous darkness, suggesting that circadian clock might be involved in the regulation of transcription of these chloroplast division related genes.