# Mitochondrial sirtuin 4 shapes the intestinal microbiota of Drosophila by controlling lysozyme expression

## TL;DR

This study shows that a protein-restricted diet increases dSirt4 in fruit flies, which affects gut bacteria by controlling lysozyme levels.

## Contribution

The study reveals a novel role of mitochondrial sirtuin 4 in regulating gut microbiota composition through lysozyme expression in Drosophila.

## Key findings

- dSirt4 knockout in intestinal cells increases lysozyme activity and alters microbiota composition.
- Lysozyme overexpression in enterocytes reduces lifespan and affects gram-positive bacteria counts.
- dSirt4-dependent lysozyme activity links dietary stress to microbiota regulation.

## Abstract

Sirtuins are deacetylases that are highly conserved throughout the animal kingdom. They act as metabolic sensors that coordinate cellular responses, allowing an adapted response to various stressors. Epithelial cells, especially those of the intestine, are directly exposed to a wide range of stressors. Together with the microbiota, they form a complex ecosystem with mutual influences. The significance of sirtuins in this complex system is still waiting to be clarified.

Here, we show that a protein-restricted diet strongly increases the intestinal expression of sirtuin 4 (dSirt4), the only mitochondrial sirtuin in Drosophila. To elucidate the effects of deregulated dSirt4 expression in the intestine, we analyzed dSirt4 knockout flies. These flies showed substantial changes in their intestinal proteome and physiological properties. One of the most striking effects was the strong induction of lysozymes in the intestine, with a corresponding increase in lysozyme activity. This effect was organ-autonomous, as it was also observed in flies with dSirt4 knocked out only in intestinal enterocytes. The significant increase in lysozyme abundance in response to tissue-specific dSirt4 knockdown did not reduce the total number of bacteria in the intestine. However, it did affect the microbiota composition by reducing the number of gram-positive bacteria. This effect on microbiota composition can be attributed to dSirt4-dependent lysozyme expression, which is absent in a lysozyme-deficient background. dSirt4 knockout in the enterocytes shortened the lifespan of the flies, as did ectopic lysozyme overexpression in the enterocytes.

The only mitochondrial sirtuin in Drosophila, dSirt4, is induced by dietary stress in intestinal epithelial cells, which directly regulates the lysozyme activity of these cells. We could associate this altered lysozyme activity with a shift in the microbiota composition, demonstrating a direct link between stress, nutrition, and the host’s microbiota regulation.

The online version contains supplementary material available at 10.1186/s42523-025-00431-x.