Renjie Shang

All Publications

• A change in<i> cis</i>-regulatory logic underlying obligate versus facultative muscle multinucleation in chordates DEVELOPMENT Johnson, C. J., Zhang, Z., Zhang, H., Shang, R., Piekarz, K. M., Bi, P., Stolfi, A. 2024; 151 (20)

  Abstract

  Vertebrates and tunicates are sister groups that share a common fusogenic factor, Myomaker (Mymk), that drives myoblast fusion and muscle multinucleation. Yet they are divergent in when and where they express Mymk. In vertebrates, all developing skeletal muscles express Mymk and are obligately multinucleated. In tunicates, Mymk is expressed only in post-metamorphic multinucleated muscles, but is absent from mononucleated larval muscles. In this study, we demonstrate that cis-regulatory sequence differences in the promoter region of Mymk underlie the different spatiotemporal patterns of its transcriptional activation in tunicates and vertebrates. Although in vertebrates myogenic regulatory factors (MRFs) such as MyoD1 alone are required and sufficient for Mymk transcription in all skeletal muscles, we show that transcription of Mymk in post-metamorphic muscles of the tunicate Ciona requires the combinatorial activity of MRF, MyoD and Early B-cell Factor (Ebf). This macroevolutionary difference appears to be encoded in cis, likely due to the presence of a putative Ebf-binding site adjacent to predicted MRF binding sites in the Ciona Mymk promoter. We further discuss how Mymk and myoblast fusion might have been regulated in the last common ancestor of tunicates and vertebrates, for which we propose two models.

  View details for DOI 10.1242/dev.202968

  View details for Web of Science ID 001345508200001

  View details for PubMedID 39114943

  View details for PubMedCentralID PMC11441980

• Loss of Myomixer Results in Defective Myoblast Fusion, Impaired Muscle Growth, and Severe Myopathy in Zebrafish. Marine biotechnology (New York, N.Y.) Wu, P., Yong, P., Zhang, Z., Xu, R., Shang, R., Shi, J., Zhang, J., Bi, P., Chen, E., Du, S. 2022; 24 (5): 1023-1038

  Abstract

  The development and growth of fish skeletal muscles require myoblast fusion to generate multinucleated myofibers. While zebrafish fast-twitch muscle can fuse to generate multinucleated fibers, the slow-twitch muscle fibers remain mononucleated in zebrafish embryos and larvae. The mechanism underlying the fiber-type-specific control of fusion remains elusive. Recent genetic studies using mice identified a long-sought fusion factor named Myomixer. To understand whether Myomixer is involved in the fiber-type specific fusion, we analyzed the transcriptional regulation of myomixer expression and characterized the muscle growth phenotype upon genetic deletion of myomixer in zebrafish. The data revealed that overexpression of Sonic Hedgehog (Shh) drastically inhibited myomixer expression and blocked myoblast fusion, recapitulating the phenotype upon direct genetic deletion of myomixer from zebrafish. The fusion defect in myomixer mutant embryos could be faithfully rescued upon re-expression of zebrafish myomixer gene or its orthologs from shark or human. Interestingly, myomixer mutant fish survived to adult stage though were notably smaller than wildtype siblings. Severe myopathy accompanied by the uncontrolled adipose infiltration was observed in both fast and slow muscle tissues of adult myomixer mutants. Collectively, our data highlight an indispensable role of myomixer gene for cell fusion during both embryonic muscle development and post-larval muscle growth.

  View details for DOI 10.1007/s10126-022-10159-3

  View details for PubMedID 36083384

  View details for PubMedCentralID PMC10112271

• Evolution of a chordate-specific mechanism for myoblast fusion. Science advances Zhang, H., Shang, R., Kim, K., Zheng, W., Johnson, C. J., Sun, L., Niu, X., Liu, L., Zhou, J., Liu, L., Zhang, Z., Uyeno, T. A., Pei, J., Fissette, S. D., Green, S. A., Samudra, S. P., Wen, J., Zhang, J., Eggenschwiler, J. T., Menke, D. B., Bronner, M. E., Grishin, N. V., Li, W., Ye, K., Zhang, Y., Stolfi, A., Bi, P. 2022; 8 (35): eadd2696

  Abstract

  Vertebrate myoblast fusion allows for multinucleated muscle fibers to compound the size and strength of mononucleated cells, but the evolution of this important process is unknown. We investigated the evolutionary origins and function of membrane-coalescing agents Myomaker and Myomixer in various groups of chordates. Here, we report that Myomaker likely arose through gene duplication in the last common ancestor of tunicates and vertebrates, while Myomixer appears to have evolved de novo in early vertebrates. Functional tests revealed a complex evolutionary history of myoblast fusion. A prevertebrate phase of muscle multinucleation driven by Myomaker was followed by the later emergence of Myomixer that enables the highly efficient fusion system of vertebrates. Evolutionary comparisons between vertebrate and nonvertebrate Myomaker revealed key structural and mechanistic insights into myoblast fusion. Thus, our findings suggest an evolutionary model of chordate fusogens and illustrate how new genes shape the emergence of novel morphogenetic traits and mechanisms.

  View details for DOI 10.1126/sciadv.add2696

  View details for PubMedID 36054355

  View details for PubMedCentralID PMC10848958

• LETMD1 is required for mitochondrial structure and thermogenic function of brown adipocytes. FASEB journal : official publication of the Federation of American Societies for Experimental Biology Snyder, M. M., Yue, F., Zhang, L., Shang, R., Qiu, J., Chen, J., Kim, K. H., Peng, Y., Oprescu, S. N., Donkin, S. S., Bi, P., Kuang, S. 2021; 35 (11): e21965

  Abstract

  Obesity and metabolic disorders caused by energy surplus pose an increasing concern within the global population. Brown adipose tissue (BAT) dissipates energy through mitochondrial non-shivering thermogenesis, thus representing a powerful agent against obesity. Here we explore the novel role of a mitochondrial outer membrane protein, LETM1-domain containing 1 (LETMD1), in BAT. We generated a knockout (Letmd1KO ) mouse model and analyzed BAT morphology, function and gene expression under various physiological conditions. While the Letmd1KO mice are born normally and have normal morphology and body weight, they lose multilocular brown adipocytes completely and have diminished mitochondrial abundance, DNA copy number, cristae structure, and thermogenic gene expression in the intrascapular BAT, associated with elevated reactive oxidative stress. In consequence, the Letmd1KO mice fail to maintain body temperature in response to acute cold exposure without food and become hypothermic within 4 h. Although the cold-exposed Letmd1KO mice can maintain body temperature in the presence of food, they cannot upregulate expression of uncoupling protein 1 (UCP1) and convert white to beige adipocytes, nor can they respond to adrenergic stimulation. These results demonstrate that LETMD1 is essential for mitochondrial structure and function, and thermogenesis of brown adipocytes.

  View details for DOI 10.1096/fj.202100597R

  View details for PubMedID 34669999

• Feedback regulation of Notch signaling and myogenesis connected by MyoD-Dll1 axis. PLoS genetics Zhang, H., Shang, R., Bi, P. 2021; 17 (8): e1009729

  Abstract

  Muscle precursor cells known as myoblasts are essential for muscle development and regeneration. Notch signaling is an ancient intercellular communication mechanism that plays prominent roles in controlling the myogenic program of myoblasts. Currently whether and how the myogenic cues feedback to refine Notch activities in these cells are largely unknown. Here, by mouse and human gene gain/loss-of-function studies, we report that MyoD directly turns on the expression of Notch-ligand gene Dll1 which activates Notch pathway to prevent precautious differentiation in neighboring myoblasts, while autonomously inhibits Notch to facilitate a myogenic program in Dll1 expressing cells. Mechanistically, we studied cis-regulatory DNA motifs underlying the MyoD-Dll1-Notch axis in vivo by characterizing myogenesis of a novel E-box deficient mouse model, as well as in human cells through CRISPR-mediated interference. These results uncovered the crucial transcriptional mechanism that mediates the reciprocal controls of Notch and myogenesis.

  View details for DOI 10.1371/journal.pgen.1009729

  View details for PubMedID 34370738

  View details for PubMedCentralID PMC8376015

• Generation of mouse conditional knockout alleles in one step using the i-GONAD method. Genome research Shang, R., Zhang, H., Bi, P. 2021; 31 (1): 121-130

  Abstract

  The Cre/loxP system is a powerful tool for gene function study in vivo. Regulated expression of Cre recombinase mediates precise deletion of genetic elements in a spatially- and temporally-controlled manner. Despite the robustness of this system, it requires a great amount of effort to create a conditional knockout model for each individual gene of interest where two loxP sites must be simultaneously inserted in cis The current undertaking involves labor-intensive embryonic stem (ES) cell-based gene targeting and tedious micromanipulations of mouse embryos. The complexity of this workflow poses formidable technical challenges, thus limiting wider applications of conditional genetics. Here, we report an alternative approach to generate mouse loxP alleles by integrating a unique design of CRISPR donor with the new oviduct electroporation technique i-GONAD. Showing the potential and simplicity of this method, we created floxed alleles for five genes in one attempt with relatively low costs and a minimal equipment setup. In addition to the conditional alleles, constitutive knockout alleles were also obtained as byproducts of these experiments. Therefore, the wider applications of i-GONAD may promote gene function studies using novel murine models.

  View details for DOI 10.1101/gr.265439.120

  View details for PubMedID 33328166

  View details for PubMedCentralID PMC7849380

• Human myotube formation is determined by MyoD-Myomixer/Myomaker axis. Science advances Zhang, H., Wen, J., Bigot, A., Chen, J., Shang, R., Mouly, V., Bi, P. 2020; 6 (51)

  Abstract

  Myoblast fusion is essential for formations of myofibers, the basic cellular and functional units of skeletal muscles. Recent genetic studies in mice identified two long-sought membrane proteins, Myomaker and Myomixer, which cooperatively drive myoblast fusion. It is unknown whether and how human muscles, with myofibers of tremendously larger size, use this mechanism to achieve multinucleations. Here, we report an interesting fusion model of human myoblasts where Myomaker is sufficient to induce low-grade fusion, while Myomixer boosts its efficiency to generate giant myotubes. By CRISPR mutagenesis and biochemical assays, we identified MyoD as the key molecular switch of fusion that is required and sufficient to initiate Myomixer and Myomaker expression. Mechanistically, we defined the E-box motifs on promoters of Myomixer and Myomaker by which MyoD induces their expression for multinucleations of human muscle cells. Together, our study uncovered the key molecular apparatus and the transcriptional control mechanism underlying human myoblast fusion.

  View details for DOI 10.1126/sciadv.abc4062

  View details for PubMedID 33355126