Abstract
The proper development, the reproduction, the response to environmental signals and even routine cellular processes of an organism depend on the proper regulation of its genome. As a consequence, cells have developed many strategies to control gene expression, ranging from the structure of chromosomes to the stability of proteins. One of the main regulators of gene activity in plants are the so-called small RNAs (sRNAs). These molecules are originated from a double stranded RNA, which is recognized and processed to 19-24 nt long sRNAs by Dicer-like (DCL) proteins. The sRNAs are then loaded into an effector complex called RISC, where they will confer specificity to the system. Gene silencing driven by sRNAs can occur both, at the transcriptional level (transcriptional gene silencing, TGS) or post-transcriptionally (post transcriptional gene silencing, PTGS). TGS is caused by DNA and/or histone methilation at the target sequence, while PTGS relies on the “slicing” of the target sequence or translation repression. sRNAs in plants can be divided in small interfering RNAs (siRNAs) or micro RNAs (miRNAs). siRNAs are originated from perfect double-stranded RNA, which can arise by different sources. On the other hand, miRNAs are processed from imperfect double-stranded RNAs that can be formed in molecules with self-complementarity. This PhD dissertation is divided in four different parts. In the first part, it is shown that random foldback structures in the genome, once transcribed, could be the source of new MIRNA genes. This scenario complements former theories, which were not sufficient to explain the origin of all new MIRNAs. In the second part of this thesis, the putative non-cell autonomy of miRNAs and trans-acting siRNAs (tasiRNA, a plant-specific class of siRNA) is investigated. By using synthetic molecules, it is shown that miRNAs can produce silencing which spread locally to a limited number of cells, while tasiRNA-triggered silencing is a long-range, cell-to-cell signal. Finally, the third and fourth parts discuss aspects related to the biogenesis of tasiRNAs. The importance and sufficiency of miR173 in starting transitivity is demonstrated and a new mechanism for gene silencing based on this finding is presented.
