Disruption of Zfh3 abolishes mulberry-specific monophagy in silkworm larvae

Feeding behavior is critical for insect survival and fitness. Most researchers have explored the molecular basis of feeding behaviors by identifying and elucidating the function of olfactory receptors (ORs) and gustatory receptors (GRs). Other types of genes, such as transcription factors, have rarely been investigated, and little is known about their potential roles. The silkworm (Bombyx mori) is a well-studied monophagic insect which primarily feeds on mulberry leaves, but the genetic basis of its monophagy is still not understood. In this report, we focused on a transcription factor encoded by the Zfh3 gene, which is highly expressed in the silkworm central and peripheral nervous systems, including brain, antenna, and maxilla. To investigate its function, Zfh3 was abrogated using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9) mutagenesis. Since Zfh3 knockout homozygotes are not viable, we studied feeding behavior in heterozygotes, and found that disruption of Zfh3 affects both gustation and olfaction. Mutant larvae lose preference for mulberry leaves, acquire the ability to consume an expanded range of diets, and exhibit improved adaptation to the M₀ artificial diet, which contains no mulberry leaves. These results provide the first demonstration that a transcription factor modulates feeding behaviors in an insect.