Abstract:The objective of this
project is to determine whether the alanine-threonine (Ala-Thr)
dipeptide found in all Myogenic Regulatory Factor (MRF) genes and known
as the "Myogenic Code", is essential for their activity. MRFs exist in
animals ranging from worms to humans, and they are typically active only
in muscle or muscle precursor cells where they play key roles in
myogenesis. MRF genes have also been shown to be involved in muscle
repair and regeneration following injury or disuse, and to play
significant roles in certain muscle abnormalities and disease states.
Thus their study has clear implications for issues of human health. This
project use~ embryos -of the ascidian Ciona intestinalis as a simple in
vivo test system to investigate the properties of MRF genes. We have two
goals. The first is to determine whether the Ala-Thr dipeptide, which is
known to be critical for the activity of vertebrate MRFs, is also
important for the myogenic activity of invertebrate MRF genes. We will
do this by making MRF proteins with mutations in this dipeptide and then
assaying the mutant proteins for the ability to stimulate muscle
development in non-muscle cells of Ciona embryos; previously we showed
that normal (i.e. non-mutant) MRFs stimulated muscle development in this
assay. Our second goal is to determine whether a potential MRF of
jellyfish - a primitive invertebrate - can direct muscle development in
our assay. A positive result would provide strong evidence that this
gene encodes an authentic MRF, which would indicate an extremely ancient
origin for this gene family. Assuming the jellyfish gene is myogenic, we
would then determine whether the Ala-Thr dipeptide it possess is
required for function. In all experiments, highly specific markers of
muscle development and differentiation are used to analyze experimental
outcomes. In addition to providing insight into the fundamental
properties of the MRF gene family all experimental procedures needed for
this project can be carried out by undergraduate students thereby
providing opportunities for their full participation in meaningful
scientific research.
