Stephen H. Devoto

Associate Professor
Ph.D. (neurobiology) The Rockefeller University

Campus Extension: 3461
Room #: Shanklin Lab 306
E-Mail: SDEVOTO@WESLEYAN.EDU

 

Muscle Fiber Type Development, Myotome Growth, and Somite Patterning

Links: 

• Devoto Lab  home page

• Publications

• What are the cellular and molecular mechanisms that lead to the development of distinct cell identities during development?

• How does Hedgehog signaling work?

• How is the correct number of each cell type determined?

What are the cellular behaviors that lead to the development of tissue form?

Our lab is addressing these broad questions by studying the development of zebrafish skeletal muscle. We are working on muscle development because muscle is a very abundant and easily accessible tissue, and also because diseases of muscle development are debilitating and common childhood diseases. We work on zebrafish because they are readily accessible for experimental manipulations throughout development and because a genetic approach to studying development is feasible in this vertebrate.

This cross section of a 24 hour old zebrafish embryo was stained with two monoclonal antibodies that each specifically label one of the two muscle fiber types. A monolayer of embryonic slow muscle fibers (stained in yellowish green) is on the surface of the myotome, surrounding the deeper fast muscle fibers (stained in red).

Muscle fibers in most vertebrates can be broadly classified as either slow or fast. Slow muscle fibers contract slowly and they fatigue slowly; in fish, slow muscle is used for steady cruising through the water. Fast muscle fibers contract fast and fatigue fast; they are used to power rapid escape responses, such as when a predator appears. Nerve activity regulates the fiber type identity of the muscle in adults; however, the earliest developing muscle fibers have an intrinsic fiber type identity. The mechanism that establishes this early fiber type identity has been completely mysterious. We are using experimental embryology and genetics, two powerful approaches that can be combined in zebrafish, to understand how the identity of these two muscle fiber types is established, and what regulates the number of each cell type (zebrafish embryonic muscle fiber types).

Our long-term goal is to understand how cell-cell interactions regulate the relative positions and proliferation of distinct skeletal muscle cell types during development. Within the trunk, two cell types, adaxial and lateral presomitic, can be unambiguously identified in the presomitic mesoderm. Adaxial cells are large, cuboidal cells adjacent to the notochord which will develop into slow muscle fibers, while lateral presomitic cells are smaller and more irregular cells in the segmental plate which will give rise to both fast muscle and sclerotome (Devoto et al., 1996). One or more members of the hedgehog (Hh) gene family triggers specification of two different types of slow muscle fibers: muscle pioneers and non-muscle pioneers (Du, et al., 1997). Finally, ectopic expression of a member of the BMP4 gene family in the notochord inhibits development of the muscle pioneer class of slow muscle fibers, without affecting development of either the non-muscle pioneer slow muscle fibers or the fast muscle fibers (Du, et al., 1997).

Our work thus suggests that the identity of a muscle fiber is determined by competing influences between a Hh gene family member and one or more members of the BMP4 gene family (our model). We are testing this model with pharmacological, molecular, and genetic approaches.

The role of Hh in embryonic slow muscle development

 Zebrafish slow muscle development offers a very powerful system in which to identify new genes involved in Hedgehog , and to test the function of known genes. We have tested whether Hh is required for slow muscle development in two ways. First, we have shown that pharmacological elimination of Hh signaling blocks the development of embryonic slow muscle fibers. Second, we have begun characterizing a new gene that we have named slow-muscle-omitted (smu). Smu mutant embryos have a nearly complete loss of slow muscle fibers. smu function is required in muscle precursors for them to respond to signaling from the notochord, and thus this gene is a good candidate for genes involved in notochord signaling. Smu is the zebrafish homologue of the Hh receptor complex protein Smoothened, and that smu is required for all Hh signal transduction. See Barresi, et al., 2000, for more information. We have also shown that all other genes which are required for Hh or midline signaling are required for slow muscle development (Stickney, et al., 2000).

Muscle growth, larval development, and fiber type identity.

It is currently not known how the number of cells in a vertebrate muscle is determined. The relatively simple zebrafish system allows us to test a number of hypotheses about the regulation of cell numbers. Twenty slow muscle fibers develop in each myotome during the embryonic period, we want to know more about what regulates the division of slow muscle precursors. To this end, we are characterizing the time course of precursor cell division, and testing whether any of the known genes are required for regulating this division.

After these initial 20 muscle fibers per somite have developed, there is a tremendous growth of the myotome, the adult zebrafish has at least 400 slow muscle fibers in each myotome. We are beginning to characterize this growth. We have recently shown that new slow muscle fibers are added beginning at about 24h of development, very shortly after embryonic slow muscle fibers form. Surprisingly, these added fibers develop independently of Hh signaling, and independently of the presence of embryonic slow muscle fibers. We are currently characterizing the development of larval fibers further.

Other aspects of somite patterning

In addition to the myotome, the somite gives rise to the vertebrae, the ribs and (in chick) the scapula. These tissues all develop from the sclerotome, which is a relatively small portion of the somite in zebrafish. We are examining the factors that regulate sclerotome development in zebrafish. Our working hypothesis is that initial sclerotome development is not dependent on midline signaling, but that sclerotome maintenance is.

What are the cellular behaviors that lead to the development of tissue form? Slow muscle precursors develop adjacent to the notochord, elongate and then migrate radially away from the notochord to become a monolayer of superficial muscle fibers (picture of the migration). This is a unique form of cell migration, and a number of questions remain unanswered about it.

Why is any of this relevant to Human Disease?

• muscular dystrophy strikes 1 in 4,000 boys in the U.S.

• heart disease accounts for over one-third of all deaths in the U.S.

• 150,000 babies are born with birth defects each year in the U.S.

• cancer accounts for one-quarter of all deaths in the U.S.
  

Our research into muscle fiber type development in zebrafish is currently funded by The Donaghue Foundation, and the National Institutes of Health.

At its most fundamental, our research is aimed at understanding why and how cells do what they do. How do they "decide" whether to divide or become specialized? How do cells decide how often to divide? How do they decide which of several possible specializations to acquire? Answers to these questions are necessary for designing appropriate and effective treatments for all of the above diseases.

More specifically, if we can understand how muscle development is triggered, and how specific types of muscle fibers are formed, then we can start to intelligently design therapies that will not only trigger muscle regeneration, but also trigger the appropriate specializations in regenerated tissues that are critical for recovery from heart disease and muscular dystrophy.