Ribosome remodeling as a mechanism of translational control during stress

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PROJECT SUMMARY Translational control is one of the major gene expression regulation mechanisms in the cell and its dysregulation leads to many human diseases. Ribosomes in general are viewed as constitutive molecular machines where protein synthesis takes place, however, this view has been recently challenged supporting the hypothesis of ribosome specialization and opening completely new field of research. This project will investigate the fundamental concept of ribosome specialization in protozoa using Leishmania as a model organism. In contrast to other eukaryotes, trypanosomatids including Leishmania species are unicellular organisms and their control of gene expression is mostly achieved during mRNA translation. Therefore, this protozoan represents an excellent model organism to study the role of ribosome specialization in mRNA translation regulation. Environment including temperature, pH, nutrition conditions plays a big role in gene expression regulation, however, it is poorly understood what molecular players are involved in the regulation of translation during environmental stresses and change of host. It is known that translation is globally repressed during the heat shock, however, some mRNAs escape translational repression and their translation is enhanced. Translation of mRNAs encoding for proteins involved in stress response is very important for the Leishmania ability to cope with stress, its differentiation and survival, however, it is not well understood how heat-induced mRNAs escape the global translational repression during the heat stress. This project is based on the hypothesis that ribosome composition undergoes a substantial change during heat stress to promote efficient translation of subset of mRNAs encoding for proteins involved in stress response. The proposed study will provide new information at several different levels: (Aim 1) it will identify on genome-wide scale subset of mRNAs that are actively translated during the heat stress; (Aim 2) it will reveal changes in ribosome composition of Leishmania during the heat stress; and finally, (Aim 3) it will examine using CRISPR/Cas9 knock-out screen what proteins indeed promote selective translation of heat-induced transcripts and what role they play in the life cycle of Leishmania. This comprehensive multidisciplinary approach will reveal for the first time how transcripts selectively rely on specific ribosome components/regulators for their efficient translation during stress in protozoa and establish their role in Leishmania differentiation.