The molecular mechanism(s) that are responsible for suppressing MyoD's transcriptional activities in undifferentiated skeletal muscle cells have not yet been determined. We now show that MyoD associates with a histone deacetylase-1 (HDAC1) in these cells and that this interaction is responsible for silencing MyoDdependent transcription of endogenous p21 as well as muscle-speci®c genes. Speci®cally, we present evidence that HDAC1 can bind directly to MyoD and use an acetylated MyoD as a substrate in vitro, whereas a mutant version of HDAC1 (H141A) can not. Furthermore, this mutant also fails to repress MyoD-mediated transcription in vivo, and unlike wild-type HDAC1 it can not inhibit myogenic conversion, as judged by confocal microscopy. Finally, we show that an endogenous MyoD can be acetylated upon its conversion to a hypophosphorylated state and only when the cells have been induced to differentiate. These results provide for a model which postulates that MyoD may be co-dependent on HDAC1 and P/CAF for temporally controlling its transcriptional activity before and after the differentiation of muscle cells.

Surface epithelial cells, such as the epidermal keratinocyte, undergo a process of terminal cell differentiation that results in the construction of a multilayered epithelium. This epithelium functions to protect the organism from the environment. Transglutaminases, enzymes that catalyze the formation of isopeptide protein-protein cross-links, are key enzymes involved in the construction of this structure. This brief review will focus on the role of these enzymes in constructing the epidermal surface.

Tazarotene-induced protein 3 (TIG3) is a recently discovered regulatory protein that is expressed in the suprabasal epidermis. In the present study, we show that TIG3 regulates keratinocyte viability and proliferation. TIG3-dependent reduction in keratinocyte viability is accompanied by a substantial increase in the number of sub-G 1 cells, nuclear shrinkage, and increased formation of cornified envelope-like structures. TIG3 localizes to the membrane fraction, and TIG3-dependent differentiation is associated with increased type I transglutaminase activity. Microscopic localization and isopeptide cross-linking studies suggest that TIG3 and type I transglutaminase co-localize in membranes. Markers of apoptosis, including caspases and poly(ADP-ribose) polymerase, are not activated by TIG3, and caspase inhibitors do not stop the TIG3-dependent reduction in cell viability. Truncation of the carboxyl-terminal membrane-anchoring domain results in a complete loss of TIG3 activity. The morphology of the TIG3-positive cells and the effects on cornified envelope formation suggest that TIG3 is an activator of terminal keratinocyte differentiation. Our studies suggest that TIG3 facilitates the terminal stages in keratinocyte differentiation via activation of type I transglutaminase. TIG31 is a novel growth regulatory protein that is a member of the H-rev107 protein family (1). This family includes Hrev107 (2), RIG1/TIG3 (1, 3), H-rev107-1 (4), H-rev107-2 (4), and A-C1 (5). Immunostaining of epidermis reveals that TIG3 is expressed in the non-proliferating, suprabasal differentiated epidermal layers, but not in the proliferative basal cells (6), suggesting that TIG3 may play an important regulatory role during keratinocyte differentiation. TIG3 levels are reduced in tumor cells, consistent with the idea that the loss of TIG3 function may contribute to disease progression in cancer (1). However, the amino acid sequence of TIG3 does not reveal any particular motif that predicts a function, and no information is available regarding the TIG3 mechanism of action. Thus, identifying the mechanism of TIG3 action is an important goal that requires knowledge regarding the effects of TIG3 on cell regulatory mechanisms. In the present study, we show that TIG3 expression in cultured keratinocytes causes a cessation of cell proliferation and enhanced cornified envelope formation. Our results suggest that TIG3 associates with membranes and that the mechanism leading to reduced cell viability involves activation of type I transglutaminase. MATERIALS AND METHODSAdenovirus Infection-The tetracycline-regulated recombinant adenovirus, tAd5-TIG3 1-164 , was constructed by cloning the TIG3 1-164 coding sequence, linked to an SV40 transcription terminator, into tAd5. This adenoviral vector contains a tetracycline-responsive element that includes the cytomegalovirus promoter and tetracycline operator. Activation of this promoter requires the presence of the tetracycline transactivator (TA) protein that is provided by co-infection with a second aden...

The epidermis is a dynamic and continually renewing surface that provides and maintains a life-sustaining interface with the environment. The epidermal keratinocyte, the major cell type of the epidermis, undergoes a complex and carefully choreographed program of differentiation. This process requires a balance between keratinocyte proliferation, differentiation, and apoptosis. This overview will concentrate on cascades that regulate the balance between keratinocyte cell proliferation and survival, and apoptosis and cell differentiation, with a particular emphasis on the role of the mitogen-activated protein kinase cascades. A summary of the literature suggests that extracellular regulated kinases function to promote keratinocyte proliferation and survival, whereas p38 mitogen-activated protein kinase functions to promote differentiation and apoptosis.

Type I transglutaminase is a plasma membrane-anchored intracellular protein-protein crosslinking enzyme that is responsible for assembly of the keratinocyte cornified envelope during terminal keratinocyte differentiation. We recently described a novel protein, TIG3, that when expressed in keratinocytes causes increased transglutaminase activity and keratinocyte cell death. However, the mechanism of activation of transglutaminase by TIG3 is not known. We now extend our previous study and show that full-length TIG3 forms a complex with type I transglutaminase that is demonstrated by TIG3-transglutaminase co-precipitation. We also demonstrate that treating TIG3-expressing cells with monodansyl cadaverine, a competitive transglutaminase substrate, attenuates the TIG3-dependent response, suggesting that transglutaminase is an important mediator of TIG3 action. These findings suggest that TIG3 forms a complex with transglutaminase resulting in transglutaminase activation and that transglutaminase activity is required for the TIG3-dependent biological response.