| Summary: | 博士 === 國立陽明大學 === 生化暨分子生物研究所 === 103 === SCUBE [signal peptide-CUB-EGF-like-containing proteins] is a novel secreted and cell membrane-associated protein family, which is composed of 3 distinctive members evolutionarily conserved in vertebrates. Scubes are expressed predominantly in a variety of developing tissues in mice, suggesting the involvement of Scubes during embryonic development. However, the developmental function of Scubes is not fully understood.
In this study, we identified and characterized zebrafish scube1 and scube3 gene, and investigated their functions during zebrafish embryonic development by using antisense morpholino (MO) knockdown approach. Whole-mount in situ hybridization revealed that zebrafish scube1 and scube3 mRNAs are maternally expressed and widely distributed during early embryonic development. Knockdown of scube1 by MO down-regulated marker genes associated with early primitive hematopoietic precursors (scl), erythroid (gata1 and hbbe1) as well as early (pu.1) and late (mpo and l-plastin) myelomonocytic lineages. Overexpression of bmp rescues the expression of scl in the posterior lateral mesoderm during early primitive hematopoietic in scube1-morphants. Biochemical and molecular analysis showed that Scube1 could interact with BMP2/4 and their receptors to augment BMP signaling. On the other hand, immunofluorescent staining of fast but not slow muscle myosin was reduced in scube3-morphants. Consistent with this finding, Scube3 knockdown in embryos specifically suppressed the expression of the myogenic transcription factor myod1 within the lateral fast muscle precursors, whereas its expression in the adaxial slow muscle precursors was largely unaffected. Additionally, we identified fgf8 as a major factor in scube3-medeated fast-muscle differentiation. In scube3-morphants, the fgf8 expression in somites was down-regulated and forced expression of fgf8 RNA restored myod1 expression in somites. Biochemical and molecular analysis showed that Scube3 may be a critical upstream regulator of fast fiber myogenesis by modulating FGF8 signaling.
In conclusion, our studies revealed the novel developmental functions of Scube gene family in primitive hematopoiesis and fast muscle development— scube1 functions at the top of the regulatory hierarchy of primitive hematopoiesis by modulating BMP activity, whereas scube3 plays an important role in fast muscle development via regulating FGF signaling.
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