Sall4 restricts glycolytic metabolism in limb buds through transcriptional regulation of glycolytic enzyme genes.
Sall4 restricts glycolytic metabolism in limb buds through transcriptional regulation of glycolytic enzyme genes.
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DOI：
10.1016/j.ydbio.2023.06.004
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发表时间：
2023-06
影响因子：
2.7
通讯作者：
H. Kawakami;Katherine Q Chen;Ruizhi Zhang;Matthew P Pappas;Abigail Bailey;J. Reisz;Dylan Corcoran;R. Nishinakamura;Angelo D’Alessandro;Y. Kawakami
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作者：
H. Kawakami;Katherine Q Chen;Ruizhi Zhang;Matthew P Pappas;Abigail Bailey;J. Reisz;Dylan Corcoran;R. Nishinakamura;Angelo D’Alessandro;Y. Kawakami
Recent studies illustrate the importance of regulation of cellular metabolism, especially glycolysis and pathways branching from glycolysis, during vertebrate embryo development. For example, glycolysis generates cellular energy ATP. Glucose carbons are also directed to the pentose phosphate pathway, which is needed to sustain anabolic processes in the rapidly growing embryos. However, our understanding of the exact status of glycolytic metabolism as well as genes that regulate glycolytic metabolism are still incomplete.Sall4is a zinc finger transcription factor that is highly expressed in undifferentiated cells in developing mouse embryos, such as blastocysts and the post-implantation epiblast.TCre; Sall4conditional knockout mouse embryos exhibit various defects in the posterior part of the body, including hindlimbs. Using transcriptomics approaches, we found that many genes encoding glycolytic enzymes are upregulated in the posterior trunk, including the hindlimb-forming region, ofSall4conditional knockout mouse embryos. In situ hybridization and qRT-PCR also confirmed upregulation of expression of several glycolytic genes in hindlimb buds. A fraction of those genes are bound by SALL4 at the promoters, gene bodies or distantly-located regions, suggesting thatSall4directly regulates expression of several glycolytic enzyme genes in hindlimb buds. To further gain insight into the metabolic status associated with the observed changes at the transcriptional level, we performed a comprehensive analysis of metabolite levels in limb buds in wild type andSall4conditional knockout embryos by high-resolution mass spectrometry. We found that the levels of metabolic intermediates of glycolysis are lower, but glycolytic end-products pyruvate and lactate did not exhibit differences inSall4conditional knockout hindlimb buds. The increased expression of glycolytic genes would have caused accelerated glycolytic flow, resulting in low levels of intermediates. This condition may have prevented intermediates from being re-directed to other pathways, such as the pentose phosphate pathway. Indeed, the change in glycolytic metabolite levels is associated with reduced levels of ATP and metabolites of the pentose phosphate pathway. To further test whether glycolysis regulates limb patterning downstream ofSall4, we conditionally inactivatedHk2, which encodes a rate-limiting enzyme gene in glycolysis and is regulated bySall4. TheTCre; Hk2conditional knockout hindlimb exhibited a short femur, and a lack of tibia and anterior digits in hindlimbs, which are defects similarly found in theTCre; Sall4conditional knockout. The similarity of skeletal defects inSall4mutants andHk2mutants suggests that regulation of glycolysis plays a role in hindlimb patterning. These data suggest thatSall4restricts glycolysis in limb buds and contributes to patterning and regulation of glucose carbon flow during development of limb buds.
