Epidermal progenitor cells are crucial in maintaining skin homeostasis and facilitating tissue replacement. The highly regulated differentiation process in epidermal progenitor cells is crucial for sustaining skin tissue integrity and preventing skin diseases, including malignancies. However, there is a wide knowledge gap regarding the molecular mechanisms regulating the differentiation process in epidermal progenitor cells. The first part of this thesis study focused on keratinocyte differentiation factor 1 (KDF1), a critical but not well studied regulator of epidermal differentiation. Through a screening assay using SILAC/MS approach, we identified and further confirmed the interaction between KDF1 and IKKα. KDF1 deficient mouse model phenotypically resembles IKKα knock-out in epidermal development. Loss of function of KDF1 enhances skin tumor growth in vivo, indicating a similar function as IKKα in tumorigenesis. Moreover, our in vitro and in vivo studies strongly supported that KDF1 and IKKα interaction is indispensable for epidermal stratification. Therefore, KDF1 and IKKα control keratinocyte differentiation by forming a regulatory complex. With further studies of the mechanism, we discovered the effect of ubiquitination on IKKα protein level and normal function during keratinocyte differentiation. We provided compelling evidence that KDF1 affect IKKα ubiquitination level by the recruitment of a deubiquitinating enzyme, USP7. In the second part of this thesis study, we performed a shRNA library screening to identify genes essential for epidermal differentiation. Vamp2 appeared on our top list, although its function in keratinocyte has never been identified. With phenotypical and histological examination of the skin from Vamp2 knock-out mouse, we hypothesized that Vamp2 protein has an essential role in regulating enucleation process during keratinocyte differentiation. Loss of Vamp2 can also enhance skin tumor growth in vivo, indicating its regulatory role in skin differentiation and maintaining tissue homeostasis. We were able to detect and visualize keratinocyte nuclear degradation both in vitro and in vivo. The subcellular localization and aggregation of Vamp2 around the nuclear membrane illustrated the potential role of Vamp2 in nuclear degradation. Via conducting SILAC-MS/MS screening, we identified various proteins associated with Vamp2 coated intracellular vesicles, including the ones involved in autophagosomes trafficking. Further functional studies are still undergoing. Taken together, our study has an important implication in the fields of skin biology as we disclosed critical and novel regulatory mechanisms controlling epidermal differentiation and skin tissue homeostasis.