Biology subjects

Datzkiw, D.

Publications and source records attributed to Datzkiw, D..

3 recordsLinked to original sources

bioRxiv2024

Wnt7a is Required for Regeneration of Dystrophic Skeletal Muscle

Intramuscular injection of Wnt7a has been shown to accelerate and augment skeletal muscle regeneration and to ameliorate dystrophic progression in mdx muscle, a model for Duchenne muscular dystrophy (DMD). However, loss-of-function studies to investigate the requirement for Wnt7a in muscle regeneration has not been evaluated. Here, we assessed muscle regeneration and function in wild type (WT) and mdx mice where Wnt7a was specifically deleted in muscle using a conditional Wnt7a floxed allele and a Myf5-Cre driver. We found that both WT and mdx mice with deletion of Wnt7a in muscle, exhibited marked deficiencies in muscle regeneration at 21 d following cardiotoxin (CTX) induced injury. Unlike WT, deletion of Wnt7a in mdx resulted in a marked decrease in specific force generation prior to CTX injury. However, both WT and mdx muscle lacking Wnt7a displayed decreased specific force generation following CTX injection. Notably the regeneration deficit observed in mdx mice lacking Wnt7a in muscle was rescued by a single tail vein injection of an extracellular vesicle preparation containing Wnt7a (Wnt7a-EVs). Therefore, we conclude that the regenerative capacity of muscle in mdx mice is due to the upregulation of endogenous Wnt7a following injury, and that systemic delivery of Wnt7a-EVs represents a therapeutic strategy for treating DMD.

genetics↗

bioRxiv2023

Wnt binding to Coatomer proteins directs secretion on exosomes independently of palmitoylation

Wnt proteins are secreted hydrophobic glycoproteins that act over long distances through poorly understood mechanisms. We discovered that Wnt7a is secreted on extracellular vesicles (EVs) following muscle injury. Structural analysis identified the motif responsible for Wnt7a secretion on EVs that we term the Exosome Binding Peptide (EBP). Addition of the EBP to an unrelated protein directed secretion on EVs. Disruption of palmitoylation, knockdown of WLS, or deletion of the N-terminal signal peptide did not affect Wnt7a secretion on purified EVs. Bio-ID analysis identified Coatomer proteins as candidates responsible for loading Wnt7a onto EVs. The crystal structure of EBP bound to the COPB2 coatomer subunit, the binding thermodynamics, and mutagenesis experiments, together demonstrate that a dilysine motif in the EBP mediates binding to COPB2. Other Wnts contain functionally analogous structural motifs. Mutation of the EBP results in a significant impairment in the ability of Wnt7a to stimulate regeneration, indicating that secretion of Wnt7a on exosomes is critical for normal regeneration in vivo. Our studies have defined the structural mechanism that mediates binding of Wnt7a to exosomes and elucidated the singularity of long-range Wnt signalling.

molecular biology↗

bioRxiv2020

N-myristoyltransferase: Tracing Steps Backwards to Find a Way Forward

N-myristoylation refers to the attachment of a 14-carbon fatty acid onto the N-terminal glycine residue of a target protein. The myristoylation reaction, catalyzed by N-myristoyltrasnferase (NMT), is essential for regulating cellular activities such as signal transduction, proliferation, migration, differentiation, and transformation. Although a considerable amount of research is performed on the overexpression of NMT in pathogenic conditions, a fundamental knowledge gap exists on the evolution of NMT and the functional impact of myristoylation for normal cellular development and functions. We performed evolutionary analyses of the NMT gene and found that most non-vertebrates harbor a single nmt gene and all vertebrates examined harbor two genes; nmt1 and nmt2. For the first time, we report that teleosts harbor two copies of nmt1, named nmt1a and nmt1b. We traced the evolutionary history of the chromosomal fragments hosting NMT1 and NMT2 in humans and found that NMT1 and NMT2 trace back to a single vertebrate ancestral chromosome. We also report the presence of putative nuclear localization sequence (NLS) and amino acid residues flanking NLS. The presence of phosphorylatable amino acid residues flanking the NLS suggests that nuclear localization of NMT is regulated by phosphorylation. The nuclear localization of NMT suggest its potential role in gene transcription.

evolutionary biology↗