Developmental Defects of Caenorhabditis elegans Lacking Branched-chain α-Ketoacid Dehydrogenase Are Mainly Caused by Monomethyl Branched-chain Fatty Acid Deficiency

Fan Jia,Minh T Than,Mingxue Cui,Min Han

doi:10.1074/jbc.m115.676650

Fan Jia, Minh T Than + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m115.676650

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Abstract

Branched-chain α-ketoacid dehydrogenase (BCKDH) catalyzes the critical step in the branched-chain amino acid (BCAA) catabolic pathway and has been the focus of extensive studies. Mutations in the complex disrupt many fundamental metabolic pathways and cause multiple human diseases including maple syrup urine disease (MSUD), autism, and other related neurological disorders. BCKDH may also be required for the synthesis of monomethyl branched-chain fatty acids (mmBCFAs) from BCAAs. The pathology of MSUD has been attributed mainly to BCAA accumulation, but the role of mmBCFA has not been evaluated. Here we show that disrupting BCKDH in Caenorhabditis elegans causes mmBCFA deficiency, in addition to BCAA accumulation. Worms with deficiency in BCKDH function manifest larval arrest and embryonic lethal phenotypes, and mmBCFA supplementation suppressed both without correcting BCAA levels. The majority of developmental defects caused by BCKDH deficiency may thus be attributed to lacking mmBCFAs in worms. Tissue-specific analysis shows that restoration of BCKDH function in multiple tissues can rescue the defects, but is especially effective in neurons. Taken together, we conclude that mmBCFA deficiency is largely responsible for the developmental defects in the worm and conceivably might also be a critical contributor to the pathology of human MSUD.

Highlights

Loss of Function of the Branched-chain ␣-ketoacid dehydrogenase (BCKDH) E2 Subunit Causes Accumulation of branched-chain amino acid (BCAA) and Reduction of monomethyl branched-chain fatty acids (mmBCFAs)—To characterize the function of BCKDH in C. elegans, we focused on the gene ZK669.4, which encodes a homologue of human dihydrolipoamide branched-chain transacylase/DBT (E2 subunit)
The ok3001 mutation deletes the entire eighth and part of the ninth exon of ZK669.4, which is expected to disrupt the catalytic domain of the encoded protein. ok3001 animals showed elevated levels of BCAAs (Fig. 1B), resembling maple syrup urine disease (MSUD) patients, and had reduced C15ISO and C17ISO levels (Fig. 1C), which are the major mmBCFAs in C. elegans [22, 39, 41]
Neither supplementation suppressed the abnormal accumulation of BCAAs in dbt-1(lf) worms. dbt1C15ISO had even further increased BCAAs in both L4 larvae and eggs (Fig. 4, B and D), which were partially viable (Fig. 3B). These results suggest that C. elegans development can tolerate elevated BCAA levels and, in contrast to what has been shown in studies on MSUD patients, mmBCFA deficiency is the major cause of the developmental phenotypes when BCKDH function is disrupted

Summary

Introduction

MmBCFA Supplementation Suppresses Larval Arrest and Embryonic Lethality Caused by dbt-1(lf)—When synchronized dbt-1(lf) L1 larvae were cultured on plates with standard food (E. coli OP50) for 3 days, 100% of them remained as L1 or L2 larvae, whereas WT worms all grew to adulthood. We noticed that, when supplemented with C17ISO, d17SPA, or lower levels of C13ISO or C15ISO, dbt-1(lf) worms grew to adults but laid only dead eggs (Fig. 3B). Supplementing worms with higher amounts of C13ISO or C15ISO, or adding C15ISO on top of C17ISO, partially rescued this phenotype (Fig. 3B), suggesting that mmBCFA deficiency, but not toxicity, causes an embryonic phenotype in dbt-1(lf) animals.

Results

Conclusion