Department of Cell and Developmental Biology / Wellstone Program
Associate Professor - University of Massachusetts Medical School
Research: Epigenetics; Muscle development and disease, FSHD; Rett Syndrome
Affiliate Investigator - Sen. Paul D. Wellstone Muscular Dystrophy Cooperative Research Center for FSHD
Principal Scientist - Boston Biomedical Research Institute
Assistant Professor - Department of Cell and Developmental Biology - University of Illinois at Urbana-Champaign
Postdoctoral fellow - Laboratory of Molecular Embryology, NICHD, NIH, Bethesda, MD. Mentors: Alan P. Wolffe and Yun-bo Shi
Research: Epigenetics, chromatin, biochemistry
Emory University, Atlanta, GA - Program in Genetics and Molecular Biology
Degree: PhD in Eukaryotic gene regulation
Miami University, Oxford, OH
Degree: B.A. in Microbiology
Epigenetic gene and genome regulation
Facioscapulohumeral muscular dystrophy (FSHD) as a model for epigenetic gene and genome regulation
All forms of FSHD are linked by epigenetic dysregulation of the 4q35 D4Z4 macrosatellite array. Many epigenetic regulatory and memory mechanisms are known to be involved in FSHD, including DNA methylation, Polycomb Group/Trithorax silencing and activation, histone modifications, chromatin remodeling, lncRNAs, sncRNAs, repeat-induced gene silencing and telomere position effect. Additional mechanisms are likely to be involved including RNA-directed DNA methylation and gene silencing, nuclear organization, and trans chromosomal interactions. Healthy individuals have an epigenetically silent chromatin structure at the 4q35 D4Z4 array whereas FSHD affected subjects have a more permissive and euchromatic structure. Thus, comparing the healthy and FSHD-affected situations provides an excellent model for investigating broadly applicable epigenetic mechanisms of gene and genome regulation.
Molecular mechanisms of FSHD pathology
FSHD is the most prevalent myopathy afflicting women, men, children and adults of all ages. FSHD is an autosomal dominant disease marked by slow but progressive atrophy in specific muscle groups in the face, upper arms, abdomen, hip girdle, and legs with individuals often showing bilateral asymmetry in muscle weakness. There is a wide range in both the age of disease onset and clinical severity, from the extremely severe infantile form (IFSHD), to the more common young adult onset FSHD manifesting in the second or third decade, to the individuals who show symptoms only much later in life, if ever. Initial symptoms often causing one to seek clinical help are difficulty raising ones arms above shoulder level, facial weakness and foot drop. Additional non-muscular symptoms that appear later in the disease progression include high-frequency hearing loss in more than half of all diagnosed cases, and vision problems in ~1% of patients. With many FSHD patients becoming wheelchair-bound as young adults, the personal, social, and economic costs of this disease are enormous, and no effective therapies exist.
FSHD is a genetic disease linked to chromosome 4q35 with a strong epigenetic component (Figure 1). FSHD research has now entered an important new stage as the DUX4 gene has emerged from studies in multiple laboratories, including our own, as the near consensus FSHD candidate gene whose misexpression is necessary for developing pathology. The DUX4gene is present in numerous copies in the genome as each 3.3kb repeat unit of a D4Z4 repeat array contains a DUX4 gene and large D4Z4 macrosatellite arrays are found in several places in the genome. However, only the DUX4 gene in the distal D4Z4 repeat of a permissive 4q35 subtelomeric array can stably express pathogenic transcripts. The current evidence supports a model in which aberrant stable expression of the DUX4 mRNA splicing variant, termed DUX4-fl (fl = full-length), in skeletal muscle is required for FSHD pathology. This pathogenic mRNA encodes a transcription factor, DUX4-FL, whose expression leads to additional aberrant expression of downstream genes in FSHD muscle. Expression of DUX4-fl per se is not necessarily causal for FSHD as healthy unaffected individuals occasionally express DUX4-fl mRNA and protein in muscle, albeit at much lower levels than seen in FSHD muscle. This suggests a quantitative model for DUX4-fl expression leading to FSHD pathology whereby low expression levels are tolerated by certain individuals and higher levels beyond a threshold result in FSHD pathology.
The primary driver of FSHD pathology is the epigenetic status of a permissive (A type subtelomere) 4q35 D4Z4 macrosatellite array. Pathogenic changes in the epigenetic status of the 4q35 D4Z4 array occur through several mechanisms including large deletions within the repeats (FSHD1), mutations in genes affecting DNA methylation and heterochromatin structure (FSHD2), or a combination of both. Regardless of the mechanism for dysregulation, these epigenetic changes result in the increased aberrant expression of DUX4-fl.
Interestingly, a number of seemingly healthy individuals have been found to possess the FSHD1 genetic lesion combined with a permissive 4q subtelomere yet show no clinical manifestation of the disease. Myogenic cells from these individuals show a wide range of DUX4-fl expression from very low, similar to healthy individuals, to levels equivalent to those found in clinically affected FSHD individuals. In addition, we have found that the epigenetic relaxation of the D4Z4 array is highly variable among genetically FSHD1 individuals, including those clinically affected as well as disease non-manifesting subjects. This indicates the existence of multiple modifier genes functioning at two levels; upstream, regulating the level of DUX4-fl expression (epigenetic modifiers) and downstream, regulating the function of DUX4-fl. Thus, in addition to DUX4-fl itself, there are a number of additional potential targets for therapeutic development.
Generation of FSHD-like model organisms
A major gap in the FSHD field is the lack of phenotypic FSHD-like model organisms based on DUX4-fl expression. This is in large part due to two large hurdles: 1) DUX4-D4Z4 is not conserved outside of old world primates, and 2) exogenous expression of DUX4-fl, even at extremely low levels, is highly cytotoxic to somatic cells of all vertebrates tested including human, mouse, Xenopus and zebrafish. However, since many of the DUX4-fl gene targets and adverse effects of expression are conserved, we believe a model organism approach can be successful and valuable. We have successfully generated DUX4-fl expressing Drosophila to use for investigating DUX4-dependent pathways. In addition, we are generating conditional lines of DUX4-fl expressing mice for developmental studies, investigating pathogenic mechanisms and for preclinical testing of potential FSHD therapeutics.
Drs. Peter and Takako Jones are a husband-wife team and function effectively as Co-PIs in the lab. We have a small but highly efficient group and have a number of productive external collaborations. Our lab has several FSHD research projects: 1) investigating the epigenetic and genetic regulation of DUX4 gene expression and alternative mRNA splicing, 2) investigating additional FSHD candidate genes and transcripts that may be involved in pathology either in concert with or independent of DUX4-fl, 3) generating animal and cell culture models of FSHD using mice, Drosophila, C. elegans and human myogenic cultures, and 4) developing therapeutic strategies to target DUX4-fl mRNA and protein expression and function.
We wish to thank the National Institute of Arthritis, Musculoskeletal and Skin Diseases, the Eunice Kennedy Shriver National Institute of Child Health and Human Development, the FSH Society, the Chris Carrino Foundation for FSHD, the Association Francaise contra les Myopathies, the Muscular Dystrophy Association and the Thoracic Foundation (Boston, MA) for their support of our research program.
Figure 1: FSHD1 and FSHD2 are linked to the epigenetic relaxation of the chromatin at a permissive chromosome 4q35 D4Z4 macrosatellite repeat array.
Each D4Z4 repeat unit encodes exons 1 and 2 of the DUX4 gene. At least one D4Z4 repeat combined with a permissive “A” type subtelomere is required to develop FSHD. The permissive “A” type subtelomere encodes a third exon containing a polyadenylation signal that is spliced onto the DUX4 mRNA, thus stabilizing only the DUX4 message transcribed from the terminal D4Z4 repeat.
*DUX4-fl mRNA and protein have been detected in a small number of healthy control biopsies and myogenic cell cultures; however, the level is significantly lower than what is found in FSHD-affected samples.
Of note, a number of individuals (estimated up to 1-3%) in the general population have FSHD1-sized deletions and do not exhibit clinical symptoms of FSHD and are referred to as non-manifesting. Their expression levels of DUX4-fl in muscle range from undetectable to levels found in FSHD-affected muscle. Interestingly, the epigenetic status of these subjects resembles that of healthy subjects as opposed to FSHD-affected. Therefore, the epigenetic status, and not the genetic status, of the FSHD-associated D4Z4 array closely correlates with disease presentation.
Department of Cell and Developmental Biology