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In the silkworm natural homozygous mutant leml, the key enzyme sepiapterin reductase (BmSPR) in the de novo synthesis pathway of BH4 is inactivated, resulting in severe deficiency of BH4 synthesis. However, it is not known why the leml larvae can survive to the second-instar stage and which pathways lead to their death when BH4 is deficient. Here, we quantified BH4 and found that the fertilized eggs contained large amounts of BH4 transferred from the mother to the offspring, maintaining its normal development in the embryo and the first instar. Subsequently, we investigated the multiple pathways in which BH4 is involved as a cofactor. The results showed that BH4 deficiency in silkworms blocked the melanin synthesis pathway, caused an insufficient degree of epidermal sclerosis, disordered tyrosine metabolism, and damaged mitochondria. On the other hand, BH4 deficiency led to the uncoupling of nitric oxide synthase (BmNOS), a reduced NO production, and a significantly reduced fat in fat body catalyzation by phospholipase A2, resulting in an impaired immune system. Meanwhile, the uncoupling of BmNOS increased the O2\u2212 content, damaged the DNA, and caused the apoptosis of the body cells. Taken together, BH4 is critical for the life and death of leml mutants. This study lays a foundation for the further exploration of lepidopteran insects and provides an important basis for the treatment of human BH4 deficiency-related diseases.<\/jats:p>","DOI":"10.3390\/ijms232012083","type":"journal-article","created":{"date-parts":[[2022,10,11]],"date-time":"2022-10-11T22:18:13Z","timestamp":1665526693000},"page":"12083","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Exploring the Mystery of the Tetrahydrobiopterin Synthetic Defect Lethal Mutant leml from Birth to Death in the Silkworm Bombyx mori"],"prefix":"10.3390","volume":"23","author":[{"given":"Dan","family":"Liang","sequence":"first","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rui","family":"Shu","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"},{"name":"Institute of Sericulture, Anhui Academy of Agricultural Sciences, 15 Huoshan Road, Hefei 230061, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Song","family":"Jiang","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"},{"name":"Anhui International Joint Research and Development Center of Sericulture Resources Utilization, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mengjun","family":"Xu","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"},{"name":"Anhui International Joint Research and Development Center of Sericulture Resources Utilization, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yangyang","family":"Cai","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hongwei","family":"Qin","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Daobo","family":"Zhang","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mengwei","family":"Feng","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Junshan","family":"Gao","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"},{"name":"Anhui International Joint Research and Development Center of Sericulture Resources Utilization, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3155-8536","authenticated-orcid":false,"given":"Yan","family":"Meng","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Anhui Agricultural University, 130 West Changjiang Road, Hefei 230036, China"},{"name":"Anhui International Joint Research and Development Center of Sericulture Resources Utilization, Hefei 230036, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,10,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1995","DOI":"10.1534\/genetics.108.096388","article-title":"Yellow and ebony Are the Responsible Genes for the Larval Color Mutants of the Silkworm Bombyx mori","volume":"180","author":"Futahashi","year":"2008","journal-title":"Genetics"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"19553","DOI":"10.1074\/jbc.M109.096743","article-title":"Mutations of an Arylalkylamine-N-acetyltransferase, Bm-iAANAT, Are Responsible for Silkworm Melanism Mutant","volume":"285","author":"Dai","year":"2010","journal-title":"J. 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