Cdc42 is a member of the Rho GTPase family of intracellular molecular switches regulating multiple signaling pathways involved in actomyosin organization and cell proliferation. Knowledge of its signaling function in mammalian cells came mostly from studies using the dominant-negative or constitutively active mutant overexpression approach in the past 2 decades. Such an approach imposes a number of experimental limitations related to specificity, dosage, and/or clonal variability. Recent studies by conditional gene targeting of cdc42 in mice have revealed its tissue-and cell type-specific role and provide definitive information of the physiological signaling functions of Cdc42 in vivo.

Lens regeneration in adult newts is a classic example of faithfully regenerating an entire organ via transdifferentiation1-6. After lentectomy, intriguing regulation allows the pigment epithelial cells (PECs) of the dorsal iris, but not the ventral, to dedifferentiate and then differentiate to form a new lens. This regulation might provide clues to the lack of lens regeneration in higher vertebrates. Six-3 and pax-6 known for their ability to induce ectopic lenses during embryogenesis7,8 and members of the BMP pathway, which are regulators of the dorsal/ventral axis establishment in embryos9 were examined for their role in induction of lens regeneration. Here we show that lens regeneration from the ventral iris is possible by inhibiting the BMP pathway or by transfecting ventral iris cells with six-3 and concomitant treatment with retinoic acid. In intact irises six-3 is expressed higher in the ventral iris. During regeneration, however, only levels in the dorsal iris are significantly increased. Such an increase is seen in ventral irises only when they are induced to transdifferentiate by six-3/RA or BMP inhibitors. Therefore, transcriptional regulation associated with competency for lens regeneration, aims to increase levels over established thresholds and not to merely render a regulatory gene as dorsal-specific. Lack of induction in the axolotl, a salamander incapable of lens regeneration seems to be associated with repression of six-3 expression.

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Intestinal stem cells (ISCs) drive small intestinal epithelial homeostasis and regeneration. Mechanistic target of rapamycin (mTOR) regulates stem and progenitor cell metabolism and is frequently dysregulated in human disease, but its physiologic functions in the mammalian small intestinal epithelium remain poorly defined. We disrupted the genes mTOR, Rptor, Rictor, or both Rptor and Rictor in mouse ISCs, progenitors, and differentiated intestinal epithelial cells (IECs) using Villin-Cre. Mutant tissues and wild-type or heterozygous littermate controls were analyzed by histologic immunostaining, immunoblots, and proliferation assays. A total of 10 Gy irradiation was used to injure the intestinal epithelium and induce subsequent crypt regeneration. We report that mTOR supports absorptive enterocytes and secretory Paneth and goblet cell function while negatively regulating chromogranin A-positive enteroendocrine cell number. Through additional Rptor, Rictor, and Rptor/Rictor mutant mouse models, we identify mechanistic target of rapamycin complex 1 as the major IEC regulatory pathway, but mechanistic target of rapamycin complex 2 also contributes to ileal villus maintenance and goblet cell size. Homeostatic adult small intestinal crypt cell proliferation, survival, and canonical wingless-int (WNT) activity are not mTOR dependent, but Olfm4(+) ISC/progenitor population maintenance and crypt regeneration postinjury require mTOR. Overall, we conclude that mTOR regulates multiple IEC lineages and promotes stem and progenitor cell activity during intestinal epithelium repair postinjury.

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