A multi-omics analysis led by Chinese scientists at Tsinghua University in Beijing has demonstrated that aberrant metabolism of the co-enzyme nicotinamide adenine dinucleotide (NAD+) was responsible for microcephaly induced by Zika virus (ZIKV) infection during pregnancy.

In addition to revealing this hitherto unknown insight into the pathogenesis of ZIKV-induced neonatal microcephaly in mice, the study identified promising new targets for its prevention and treatment, the authors reported in the August 12, 2021, edition of Nature Metabolism.

ZIKV is a flavivirus related to the dengue, West Nile and yellow fever viruses, which has become a major public health challenge in the subtropical areas of central America and southeast Asia endemic for the ZKIV Aedes mosquito.

While postnatal infections are typically mild, ZIKV infection during pregnancy can induce brain cell death and neonatal microcephaly, but remains untreatable.

Therefore, it is important to improve our understanding of ZIKV-induced microcephaly pathogenesis, in order to develop new therapies.

Viruses have evolved multiple strategies to reprogram the metabolism of infected host cells, enabling rapid viral replication and escape from host immune responses.

Identifying such viral-induced host cell metabolic reprograming could contribute to a better understanding of virus-host interactions and help to develop potential antiviral strategies by targeting metabolic alterations.

However, little is known about the in vivo impacts of ZIKV infection on host metabolism and pathogenesis of microcephaly, which can be revealed by multi-omics analysis.

Multi-omics analysis

Multi-omics analysis allows for comprehensive understanding of the molecular mechanisms underlying complex diseases, including cancer, diabetes and viral infections.

"Our multi-omics analysis comprised transcriptomics, proteomics, phosphoproteomics and metabolomics, which is more informative than conventional genomic analyses," said study leader Zeping Hu, a professor in the School of Pharmaceutical Sciences at Tsinghua University.

"Not all diseases are caused by genetic defects, of which acute viral infectious diseases are perfect examples," Hu told BioWorld Science.

"Integrated analyses of diverse molecules including DNA, RNA, protein and metabolites may provide more comprehensive information on biological pathways and enable an improved understanding of both genotype and phenotype, together with regulatory crosstalk at different molecular levels."

Transcriptomics studies in cell lines and animal models have shown that ZIKV infection causes strong immune responses and aberrant neurodevelopment, but to date there have been no in vivo integrative analyses of ZIKV-induced microcephaly using multi-omics techniques.

Consequently, in the new Nature Metabolism study, Hu and his research team performed an extensive and systematic multi-omics analysis of brain tissues from mock- and ZIKV-infected mouse brains.

Proteomics and metabolomics analyses uncovered marked alterations in NAD+-related metabolic pathways, including oxidative phosphorylation (OXPHOS), the tricarboxylic acid (TCA) cycle and tryptophan metabolism.

"Reduced NAD+ levels and downregulation of OXPHOS and the TCA cycle were seen in ZIKV-infected brains, emphasizing the possibility that NAD+ depletion caused bioenergy deficiency and that mitochondrial dysfunction may contribute to cell death in ZIKV-induced microcephaly," said Hu.

Moreover, the phosphoproteomics analysis indicated that mitogen-activated protein kinase (MAPK) and cyclic GMP-protein kinase G (cGMP-PKG) signaling might be associated with ZIKV-induced microcephaly.

"Enrichment analysis of these differential phosphoproteins allowed identification of specific phosphorylation events occurring throughout a given cellular pathway, focusing our attention on the apparently strongly upregulated cGMP-PKG and MAPK signaling pathways," explained Hu.

"MAPK signaling was also found to regulate NAD+ metabolism via nicotinamide nucleotide adenyltransferase 2 (NMNAT2), thereby promoting axon degeneration," he noted.

"Given that NAD+ was reduced, that NMNAT2 was downregulated at the mRNA and protein levels, and that MAPK signaling was upregulated in ZIKV-infected brains, we speculated that the MAPK-NMNAT2-NAD+ axis may have a crucial neuron-specific impact on the pathogenesis of ZIKV-induced microcephaly."

Notably, the researchers demonstrated the usefulness of these multi-omics datasets with follow-up in vivo experiments.

"We injected NAD+ into the brain and provided direct evidence that such NAD+ supplementation showed significant protective effects on ZIKV-induced cell death and cortex thinning," Hu said.

Similarly, "systemic NR supplementation significantly reduced cell death induced by ZIKV infection in mouse brains and, importantly, markedly increased cortical thickness," he said. "Moreover, we showed that NR treatment increased brain weight and body weight, and significantly improved survival of ZIKV-infected mice, revealing that NR confers protective effects against ZIKV-induced microcephaly at both the molecular and systemic levels."

Collectively, these findings support future investigations of ZIKV-induced microcephaly and suggest that metabolic alterations might be exploited for developing therapeutic strategies for ZIKV and possibly other virus-induced diseases.

"Our study has provided rich datasets to facilitate a deeper mechanistic understanding of the cellular and molecular basis of ZIKV-induced microcephaly, uncovering multiple attractive therapeutic targets," noted Hu.

"Importantly, this NAD+ boosting strategy showing promise in mice needs to be tested in human preclinical trials, either alone or in combination with other antiviral therapy, which our group hope to investigate in the near future."