Translation has emerged as a new and important layer of regulation at the last step in the journey from gene to gene product. This regulation is conferred in part through the activity of specialized ribosomes which vary in their complement of ribosomal proteins and have the capacity to selectively translate specific subsets of mRNAs. While this mechanism may offer cells an unparalleled ability to rapidly adjust protein output, the extent and impact of ribosome-mediated translational regulation has not yet been addressed in terminally differentiated cells and tissues or at the organismal level. My preliminary studies demonstrate that translational regulation may be of particular importance in promoting tissue regeneration in the axolotl - a species of highly regenerative salamander. Using axolotl limb amputation as a model, I demonstrate that unlike in mammals, severe injury in this species triggers a rapid translational response which selectively targets translation of ribosomal proteins. In this proposal I examine the role of regulated protein synthesis in the axolotl and mouse; I establish the role of specialized ribosomes in the process of axolotl limb regeneration and I extend the scope of this research to murine chondrocytes and a novel human cartilage organoid system, in order to define the role of specialized ribosomes in the development, regeneration and repair of vertebrate tissues. The long-term goal of this work is to utilize in vitro and in vivo models, including micromass murine chondrocyte culture and human cartilage organoids, genetic modeling in mice and regeneration modeling in axolotls, coupled with state-of-the-art mass spectrometry and ribosome/polysome profiling methods to define the role of ribosome-mediated translational regulation in tissue development and disease and to identify new genes and factors with therapeutic potential in tissue regeneration, with a particular emphasis on cartilage health. The research proposed in this application will be carried out within the highly collaborative environment of Stanford University and supported by a multidisciplinary advisory committee with expertise in translation control, glycobiology, axolotl regeneration and stem cell differentiation. Upon completion of the K99 phase, the candidate?s goals are to continue this work as an independent investigator in an academic research setting.
Specialized ribosomes act as sophisticated molecular gatekeepers that regulate when, where and which proteins are made. Using axolotl, mouse, and in vitro human culture systems, I investigate the role of ribosome-mediated control of protein synthesis in the context of limb regeneration in the axolotl and in the development and differentiation of mammalian cartilage. This work defines the impact of translation as a new layer of gene regulation in the organismal context.
