Transgenic Ectopic Overexpression of Broad Complex (BrC-Z2) in the Silk Gland Inhibits the Expression of Silk Fibroin Genes of Bombyx mori

Abstract

Bombyx mori silk protein genes are strictly turned on and off in different developmental stages under the hormone periodically change. The broad complex (BrC) is a transcription factor mediating 20-hydroxyecdysone action, which plays important roles during metamorphosis. Here, we observed that two isoforms of BmBrC (BmBrC-Z2 and BmBrC-Z4) exhibited contrasting expression patterns with fibroin genes (FibH, FibL and P25) in the posterior silk gland (PSG), suggesting that BmBrC may negatively regulate fibroin genes. Transgenic lines were constructed to ectopically overexpress BmBrC-Z2 in the PSG. The silk protein genes in the transgenic line were decreased to almost half of that in the wild type. The silk yield was decreased significantly. In addition, the expression levels of regulatory factors (BmKr-h1 and BmDimm) response to juvenile hormone (JH) signal were inhibited significantly. Then exogenous JH in the BmBrC-Z2 overexpressed lines can inhibit the expression of BmBrC-Z2 and activate the expression of silk protein genes and restore the silk yield to the level of the wild type. These results indicated that BmBrC may inhibit fibroin genes by repressing the JH signal pathway, which would assist in deciphering the comprehensive regulation mechanism of silk protein genes.

Dynamics of Nuclear Matrix Attachment Regions During 5th Instar Posterior Silk Gland Development in Bombyx Mori

“…Our data suggest that the MARs found in the coding regions of Br-c may be involved in the down-regulation of the broad-complex which in turn increase the FIBH expression. This is consistent with studies that showed ectopic expression of Br-c Z2 inhibited FIBH expression (67).…”

“…Previous work from our lab showed that the broad-complex negatively regulates broin gene expression (66).This was further supported by a study by a recent study that showed that the overexpression of BmBr-c Z2 in the transgenic lines of B. mori led to decline in the FIBH, Fib-l and P25 silk proteins. This mechanism was shown to be connected with the juvenile hormone pathway which is linked to the expression of its transcription factors, Bmdimm and BmKr-h1 (67). The expression of Bmdimm was shown to positively regulate FIBH which correlates with the effect of Br-c expression on the regulation of FIBH expression (68).…”

Dimmed gene knockout shortens larval growth and reduces silk yield in the silkworm, Bombyx mori

“…Generally, silkworm development is affected by changes in the prothoracic gland, corpus allatum, fat body, hemolymph, and other organs; however, abnormal silk protein synthesis and silk gland development did not affect the development of larvae (Cong et al, 2020; Hu et al, 2019; Ma et al, 2014). Therefore, we speculate that Bmdimm may also function in other organs.…”

“…mori ‐derived salivary gland‐expressed bHLH (Bmsage) (Zhao et al, 2014), B . mori ‐derived dimmed (Bmdimm) (Zhao et al, 2015); negative regulatory factors, fibroin modulator‐binding protein 1 (FMBP‐1) (Liu et al, 2019; Takiya et al, 1997), fushi tarazu‐transcription factor 1 (Ftz‐F1) (Zhou, Zha, Shi, Zhao, et al, 2016), multiprotein bridging factor 2 (MBF2) (Zhou, Zha, Shi, Wei, et al, 2016), broad complex isoform 2 (Brc‐Z2) (Cong et al, 2020); and silk gland factor‐4 (SGF4) (Hui et al, 1990) and fem‐3 binding factor (FBF‐A1) (Hui et al, 1990). Most transcription factors, including the Bmdimm gene, are involved in the regulation of FibH.…”

“…Several transcription factors are involved in the regulation of fibroin and sericin genes, including positive regulatory factors, silk gland factor 1/fork head (SGF1/Fkh) (Mach et al, 1995;Zhao et al, 2014), silk gland factor 2 (SGF2) (Kimoto et al, 2015;Ohno et al, 2013), silk gland factor 3/POU-domain protein (SGF3/POU-M1) (Kimoto et al, 2012;Xu et al, 1994) , B. mori-derived salivary gland-expressed bHLH (Bmsage) (Zhao et al, 2014), B. mori-derived dimmed (Bmdimm) (Zhao et al, 2015); negative regulatory factors, fibroin modulator-binding protein 1 (FMBP-1) (Liu et al, 2019;Takiya et al, 1997), fushi tarazutranscription factor 1 (Ftz-F1) (Zhou, Zha, Shi, Zhao, et al, 2016), multiprotein bridging factor 2 (MBF2) (Zhou, Zha, Shi, Wei, et al, 2016), broad complex isoform 2 (Brc-Z2) (Cong et al, 2020); and silk gland factor-4 (SGF4) (Hui et al, 1990) and fem-3 binding factor (FBF-A1) (Hui et al, 1990). Most transcription factors, including the Bmdimm gene, are involved in the regulation of FibH.…”