Combining Fluorescent In Situ Hybridization with Immunofluorescence and Lectin Staining in Planarians.

The capability to simultaneously apply different molecular tools to visualize a wide variety of changes in genetic expression and tissue composition in Schmidtea mediterranea has always been of great interest. The most commonly used techniques are fluorescent in situ hybridization (FISH) and immunofluorescence (IF) detection. Here, we describe a novel way to perform both protocols together adding the possibility to combine them with fluorescent-conjugated lectin staining to further broaden the detection of tissues. We also present a novel lectin fixation protocol to enhance the signal, which could be useful when single-cell resolution is required.

Planarian cell number depends on blitzschnell, a novel gene family that balances cell proliferation and cell death.

Control of cell number is crucial to define body size during animal development and to restrict tumoral transformation. The cell number is determined by the balance between cell proliferation and cell death. Although many genes are known to regulate those processes, the molecular mechanisms underlying the relationship between cell number and body size remain poorly understood. This relationship can be better understood by studying planarians, flatworms that continuously change their body size according to nutrient availability. We identified a novel gene family, blitzschnell (bls), that consists of de novo and taxonomically restricted genes that control cell proliferation:cell death ratio. Their silencing promotes faster regeneration and increases cell number during homeostasis. Importantly, this increase in cell number leads to an increase in body size only in a nutrient-rich environment; in starved planarians, silencing results in a decrease in cell size and cell accumulation that ultimately produces overgrowths. bls expression is downregulated after feeding and is related to activity of the insulin/Akt/mTOR network, suggesting that the bls family evolved in planarians as an additional mechanism for restricting cell number in nutrient-fluctuating environments.