Key points are not available for this paper at this time.
Regulated proteolysis has been postulated to be critical for proper control of cell functions. Muscle development, in particular, involves a great deal of structural adaptation and remodeling mediated by proteases. The transcription factor YY1 represses muscle-restricted expression of the sarcomeric α-actin genes. Consistent with this repressor function of YY1, the nuclear regulator is down-regulated at the protein level during skeletal as well as cardiac muscle cell differentiation. However, the YY1 message remains relatively unaltered throughout the myoblast-myotube transition, implicating a post-translational regulatory mechanism. We show that YY1 can be a substrate for cleavage by the calcium-activated neutral protease calpain II (m-calpain) and the 26 S proteasome. The calcium ionophore A23187 destabilized YY1 in cultured myoblasts, and the decrease in YY1 protein levels could be prevented by calpain inhibitor II and calpeptin. Treatment with the proteasome inhibitors MG132 and lactacystin resulted in the stabilization of YY1 protein, which is consistent with the finding that YY1 is readily polyubiquitinated in reticulocyte lysates. We further show that proteolytic targeting by calpain II and the proteasome involves different structural elements of YY1. This study thus illustrates two proteolytic pathways through which the transcriptional regulator can be differentially targeted under different cell growth conditions. Regulated proteolysis has been postulated to be critical for proper control of cell functions. Muscle development, in particular, involves a great deal of structural adaptation and remodeling mediated by proteases. The transcription factor YY1 represses muscle-restricted expression of the sarcomeric α-actin genes. Consistent with this repressor function of YY1, the nuclear regulator is down-regulated at the protein level during skeletal as well as cardiac muscle cell differentiation. However, the YY1 message remains relatively unaltered throughout the myoblast-myotube transition, implicating a post-translational regulatory mechanism. We show that YY1 can be a substrate for cleavage by the calcium-activated neutral protease calpain II (m-calpain) and the 26 S proteasome. The calcium ionophore A23187 destabilized YY1 in cultured myoblasts, and the decrease in YY1 protein levels could be prevented by calpain inhibitor II and calpeptin. Treatment with the proteasome inhibitors MG132 and lactacystin resulted in the stabilization of YY1 protein, which is consistent with the finding that YY1 is readily polyubiquitinated in reticulocyte lysates. We further show that proteolytic targeting by calpain II and the proteasome involves different structural elements of YY1. This study thus illustrates two proteolytic pathways through which the transcriptional regulator can be differentially targeted under different cell growth conditions. Many cellular processes are known to be controlled by short-lived proteins, including products of the proto-oncogenes, cell cycle regulators, and developmentally regulated transcription factors (1Carillo S. Pariat M. Steff A.M. Roux P. Etienne-Julan M. Lorca T. Piechaczyk M. Oncogene. 1994; 9: 1679-1689PubMed Google Scholar, 2Hochstrasser M. Ellison M.J. Chau V. Varshavsky A. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 4606-4610Crossref PubMed Scopus (208) Google Scholar, 3Yaglom J. Linskens M.H.K. Sadis S. Rubin D.M. Futcher B. Finley D. Mol. Cell. Biol. 1995; 15: 731-741Crossref PubMed Google Scholar). The fast turnover of these regulatory proteins reflects a metabolic requirement for rapidly changing their concentrations and is presumably mediated by a complex interplay among various proteases and protease inhibitors. Selective degradation of transcriptional activators and repressors, in particular, may provide efficient regulatory mechanisms contributing to the rapid shut-off and turn-on of gene activity, respectively. The recent study of the pleiotropic transcription factor NF-κB has revealed that the ubiquitin-proteasome pathway can function not only in the complete degradation of proteins but also in the regulated processing of precursors into active transcription factors (4Palombella V.J. Rando O.J. Goldberg A.L. Maniatis T. Cell. 1994; 78: 773-785Abstract Full Text PDF PubMed Scopus (1935) Google Scholar, 5Hershko A. Ciechanover A. Annu. Rev. Biochem. 1992; 61: 761-807Crossref PubMed Scopus (1226) Google Scholar). Calpains represent the other major class of nonlysosomal proteases functioning in a calcium-dependent fashion (6Goll D.E. Thompson V.F. Taylor R.G. Christiansen J.A. Biochimie (Paris). 1992; 74: 225-237Crossref PubMed Scopus (219) Google Scholar, 7Suzuki K. Saido T.C. Hirai S. Ann. N. Y. Acad. Sci. 1992; 674: 218-227Crossref PubMed Scopus (112) Google Scholar). Interestingly, both the proteasome and calpains have been found to play a regulatory role in the function and/or stability of c-Fos and the tumor suppressor protein p53 (8Stancovski I. Gonen H. Orian A. Schwartz A.L. Ciechanover A. Mol. Cell. Biol. 1995; 15: 7106-7116Crossref PubMed Scopus (159) Google Scholar, 9Hirai S. Kawasaki H. Yaniv M. Suzuki K. FEBS Lett. 1991; 287: 57-61Crossref PubMed Scopus (150) Google Scholar, 10Maki C.G. Huibregtse J.M. Howley P.M. Cancer Res. 1996; 56: 2649-2654PubMed Google Scholar, 11Kubbutat M.H.G. Vousden K.H. Mol. Cell. Biol. 1997; 17: 460-468Crossref PubMed Scopus (280) Google Scholar). Thus, rapid degradation or processing of specific transcription factors can underlie a wide range of dynamic cellular and developmental processes. Muscle development involves a great deal of structural adaptation and remodeling mediated by induced protein synthesis and degradation (12Dayton W.R. Schollmeyer J.V. Lepley R.A. Cortes L.R. Biochim. Biophys. Acta. 1981; 659: 48-61Crossref PubMed Scopus (173) Google Scholar, 13Couch C.B. Strittmatter W.J. Cell. 1983; 32: 257-265Abstract Full Text PDF PubMed Scopus (96) Google Scholar, 14Kumar A. Shafiq S. Wadgaonkar R. Stracher A. Cell. Mol. Biol. 1992; 38: 687-691PubMed Google Scholar). Although protein turnover must be highly selective if it is to be developmentally useful, little is known concerning the regulatory mechanisms responsible for protein targeting and subsequent degradation during development. The expression of a lysosomal cysteine protease family, cathepsins, was found to increase during muscle differentiation (15Jane D.T. Dufresne M.J. Biochem. Cell Biol. 1994; 72: 267-274Crossref PubMed Scopus (13) Google Scholar). Myoblast fusions in chick and rats were both shown to require metalloendoprotease activity (16Hayward L.J. Schwartz R.J. J. Cell Biol. 1986; 102: 1485-1493Crossref PubMed Scopus (112) Google Scholar). Consistent with the observed calcium influx during myoblast membrane fusion (17David J.D. See W.M. Higginbotham C. Dev. Biol. 1981; 82: 297-307Crossref PubMed Scopus (88) Google Scholar), the activity of the calcium-activated neutral protease (calpain) is up-regulated during and required for myogenesis (14Kumar A. Shafiq S. Wadgaonkar R. Stracher A. Cell. Mol. Biol. 1992; 38: 687-691PubMed Google Scholar, 18Kwak K.B. Chung S.S. Kim O.M. Kang M.S. Ha D.B. Chung C.H. Biochim. Biophys. Acta. 1993; 1175: 243-249Crossref PubMed Scopus (72) Google Scholar). These findings suggest that temporal regulation of proteolytic events plays an important role in muscle development. In most cases, however, the endogenous protein substrates proteolyzed during differentiation have not been characterized, and the physiological relevance remains to be examined. We have previously shown that YY1, a C2H2-type zinc finger DNA-binding protein (19Shi Y. Seto E. Chang L.S. Shenk T. Cell. 1991; 67: 377-388Abstract Full Text PDF PubMed Scopus (847) Google Scholar), is capable of simultaneously activating and repressing the expression of the c-mycproto-oncogene and the sarcomeric α-actin gene, respectively (20Lee T.C. Zhang Y. Schwartz R.J. Oncogene. 1994; 9: 1047-1052PubMed Google Scholar). In chick embryonic myoblast culture, YY1 inhibits muscle-restricted transcription of the skeletal α-actin gene by excluding SRF, a positive MADS-box myogenic transcription factor, from the most proximal serum response element of the actin gene promoter (20Lee T.C. Zhang Y. Schwartz R.J. Oncogene. 1994; 9: 1047-1052PubMed Google Scholar, 21Lee T.C. Chow K.L. Fang P. Schwartz R.J. Mol. Cell. Biol. 1991; 11: 5090-5100Crossref PubMed Google Scholar). In these studies, myoblasts rendered incapable of differentiation were found to contain higher levels of YY1 and c-Myc proteins compared with the differentiated myotubes. Given the established inhibitory effect of c-myc on myogenic differentiation (22Miner J.H. Wold B.J. Mol. Cell. Biol. 1991; 11: 2842-2851Crossref PubMed Scopus (153) Google Scholar) and the activating effect of YY1 on c-myc (20Lee T.C. Zhang Y. Schwartz R.J. Oncogene. 1994; 9: 1047-1052PubMed Google Scholar, 23Riggs K. Saleque S. Wong K.K. Merrell K.T. Lee J.S. Shi Y. Calame K. Mol. Cell. Biol. 1993; 13: 7487-7495Crossref PubMed Scopus (168) Google Scholar), the down-regulation of YY1 was postulated to be essential for the expression of the sarcomeric α-actin genes. However, the YY1 down-regulation mechanism during myogenesis remains unclear. Here we report that YY1 is similarly down-regulated during the in vitro differentiation of cultured rat skeletal myoblasts and ventricular cardiomyocytes. In contrast to the down-regulation of YY1 protein, the YY1 message remains unaltered throughout the myoblast-myotube transition. We present both in vivo and in vitro evidence that calpains and the 26 S proteasome are involved in the in vivo stability of YY1. This finding illustrates a post-translational mechanism through which the repressor of myogenic transcription may be selectively inactivated by developmentally regulated proteolysis to facilitate muscle development. Primary skeletal and cardiac muscle cell cultures were prepared as described (24Kalenik J.L. Chen D. Bradley M.E. Chen S.J. Lee T.C. Nucleic Acids Res. 1997; 25: 843-849Crossref PubMed Scopus (87) Google Scholar, 25Chen S.J. Bradley M.E. Lee T.C. Mol. Cell. Biochem. 1998; (in press)Google Scholar). In brief, minced tissues were gently agitated in 15 ml of 0.05% trypsin + 1 mm EDTA at 4 °C overnight. Excessive trypsin solution was removed following overnight agitation, and tissue fragments further incubated at 37 °C for 10 min, after which 10 ml of minimal essential medium (MEM) 1The abbreviations used are: MEM, minimal essential medium; PAGE, polyacrylamide gel electrophoresis; TLCK, tosyl-l-lysine chloromethyl ketone. containing 10% horse serum was added to inactivate trypsin. Collagenase (Worthington) and DNase I (Sigma) were then added to a final concentration of 1 mg/ml and 0.1 mg/ml, respectively, for another 20 min of incubation. Tissue fragments were then triturated three to four cycles with the addition of 10 ml of medium for each cycle. Skeletal myoblasts were plated in MEM + 10% fetal bovine serum on Primaria culture dishes (Falcon) at a density of 3,120 cells/mm2. Cardiac myocytes were plated in MEM + 10% horse serum at a density of 1,040 cells/mm2. Cultured mouse C2 and Sol8 myoblasts were maintained in MEM + 10% fetal bovine serum and 50 μg/ml gentamicin. Calpain inhibitor II and calpeptin were purchased from Calbiochem. MG132 was provided by Cecile Pickart (Johns was purchased from E. J. 10 Sol8 were by in and were and The cell was in ml of mm mm 0.1 10% and 1 mm were at 4 °C by in a of 20 The cell was and the was and in at This protein was used as the of endogenous The in vitro cleavage was in a of 20 In each of YY1 was used as Sol8 myoblast or calpain II from or was used as the of were on and was added to the cleavage were at 37 °C for min for or 20 min for calpain were with with were for min, by and then by the of protease a was used M. Biochem. 1993; PubMed Scopus Google Scholar). were in 15 containing 10 mm mm 0.1 mm mm and of control was also for each protease were by various protease inhibitors and 1 of calpain II and to at for of was added to the and of was with ml of protein was by a Calpain activity was as decrease in in the of In of YY1 was reticulocyte of purchased from were in containing of reticulocyte and 0.1 of YY1 and incubated at 37 °C for were then for 10% and an as described was the was in with concentration 0.1 and on a gel containing was to membrane and by was in solution 10% and 1 for and then incubated with a YY1 at °C overnight. membrane was with at °C by with was and for were by at the and in with 1 mm and were for were to The YY1 and were described previously (24Kalenik J.L. Chen D. Bradley M.E. Chen S.J. Lee T.C. Nucleic Acids Res. 1997; 25: 843-849Crossref PubMed Scopus (87) Google Scholar). The was used to an embryonic muscle that myoblasts contain higher levels of YY1 protein but levels of protein differentiated myotubes. YY1 and are also differentially regulated in we muscle cell cultures prepared from shown in that the YY1 protein levels were the in rat myoblasts and were and myoblasts were through cell is that the YY1 protein after was from we found that contain higher levels of YY1 T.C. Chow K.L. Fang P. Schwartz R.J. Mol. Cell. Biol. 1991; 11: 5090-5100Crossref PubMed Google Scholar). 1 that protein levels were during myogenesis and were the in differentiated The developmental of YY1 protein was further in cultured rat ventricular cardiac myocytes The study that YY1 was similarly down-regulated during in vitro cardiac differentiation. Cardiac maintained under the differentiation medium YY1 protein a down-regulation mechanism to muscle cell of YY1 protein during ventricular cardiac differentiation. Cardiac were from were to and plated in MEM containing 10% fetal bovine serum and 20 were maintained in the growth medium for in the of after which culture medium was to the differentiation medium were at the and were for cardiac proteins with YY1 cardiac proteins with cardiac proteins with YY1 were maintained in differentiation medium as Although YY1 is to be in tissues it may a a regulatory show that YY1 protein is down-regulated during skeletal as well as ventricular cardiac muscle cell differentiation. in YY1 protein could be at we to the of YY1 The temporal of YY1 expression during skeletal muscle cell differentiation was by in which that YY1 levels not from myoblasts to Consistent with the YY1 levels were found to relatively during cell differentiation E. K. Mol. Cell. Biol. 1992; PubMed Scopus Google Scholar). Thus, the down-regulation of YY1 during muscle cell differentiation is most at the protein The findings to YY1 be down-regulated by a mechanism during Calpains were in skeletal muscle PubMed Scopus (173) Google Scholar), and activity was found to be up-regulated and essential during myogenic differentiation. is the down-regulation of YY1 protein during myoblast fusion which has been known to be mediated by calcium influx (17David J.D. See W.M. Higginbotham C. Dev. Biol. 1981; 82: 297-307Crossref PubMed Scopus (88) Google Scholar). the of the in the degradation of YY1, we an in vitro calcium-dependent proteolysis myoblast as the of endogenous YY1 and degradation were by YY1 4 that degradation of YY1 was of YY1 with the or with the addition of of 1 mm on the other resulted in the of a cleavage by the YY1 This finding that YY1 may be a substrate of endogenous calpain II which levels of calcium for activity as to calpain I which levels of calcium We further the calpain 4 the of the YY1 cleavage by the calpain II in the of 1 calpain II cleavage of YY1 in the of or at We on to YY1 protein be in with specific calpain inhibitors. Primary myoblasts were used to the effect of two specific calpain calpeptin and calpain inhibitor II M.E. N. S. J. 1994; PubMed Scopus Google Scholar), on the level of YY1. However, the myoblasts were found to be to the and the resulted in a of not This finding presumably a role for calpains in the of cultured Sol8 myoblasts C. C. P. J. J. D. J. 1991; PubMed Scopus Google Scholar) were thus used in the subsequent inhibitor of protease calpain inhibitor II M.E. N. S. J. 1994; PubMed Scopus Google Scholar), of J. J.L. J.A. J. Biochem. 1986; PubMed Scopus Google Scholar), and MG132 of the 26 S Goldberg A.L. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar) were for their inhibitory on calpain II activity a in described previously M. Biochem. 1993; PubMed Scopus Google Scholar). that not calpain II activity as calpain inhibitor II and which are of and respectively, each a of the activity of calpain Thus, MG132 to be capable of the of both calpains and the proteasome. We on to the inhibitors YY1 protein in the Sol8 myoblasts were with the and were to the effect on YY1 protein concentrations for the were that minimal of cell was observed after that YY1 protein levels were not by or calpain inhibitor II of Sol8 myoblasts with MG132 YY1 protein at or with higher of MG132 to a rapid of myoblast not and activity of calpains require calcium levels in myoblasts, an with myoblast fusion during myogenesis (17David J.D. See W.M. Higginbotham C. Dev. Biol. 1981; 82: 297-307Crossref PubMed Scopus (88) Google Scholar), an effect of the calpain inhibitor on YY1 protein stability may require calcium levels in the provide evidence this calpains were by myoblasts with the calcium ionophore which has been shown to myogenesis (14Kumar A. Shafiq S. Wadgaonkar R. Stracher A. Cell. Mol. Biol. 1992; 38: 687-691PubMed Google Scholar) and calpains J. E. J. Biol. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar). that YY1 protein levels were by A23187 in Sol8 Calpain inhibitor II as well as MG132 were both to YY1 in Sol8 these in vitro and in vivo suggest the of the in proteolysis of YY1. a well known inhibitor of the 26 S was to YY1 in myoblasts that YY1 is to be a substrate of the proteasome as However, the shown with of M.E. N. S. J. 1994; PubMed Scopus Google Scholar, K.L. C. D. R. D. Goldberg A.L. Cell. 1994; 78: Full Text PDF PubMed Scopus Google Scholar) that MG132 can both calpains and the proteasome. this we further the effect of which is highly specific for the proteasome Proc. Natl. Acad. Sci. U. S. A. 1994; PubMed Scopus Google Scholar). We have shown previously that a polyubiquitinated YY1 which could not be in myoblasts, could be in myoblasts of serum L.J. Chen S.J. J.L. Bradley M.E. Lee T.C. Cell Res. 1996; PubMed Scopus Google Scholar). Thus, myoblasts maintained in growth medium as well as medium were with lactacystin for and YY1 protein levels were by that lactacystin not have a effect on the stability of YY1 in myoblasts it YY1 protein levels in myoblasts further provide evidence that YY1 is a we an in reticulocyte as an established of A. Ciechanover A. Annu. Rev. Biochem. 1992; 61: 761-807Crossref PubMed Scopus (1226) Google Scholar). In the in YY1 was incubated with reticulocyte and products were by and with YY1 of protein to the YY1 protein could be which to YY1 proteins to or The YY1 on the other was not by as by the of We thus that the zinc finger of YY1 contain for the proteolytic is readily in reticulocyte lysates. 0.1 of YY1 and the were each incubated with reticulocyte as described in the were by 10% by YY1 The of the YY1 protein is by the of YY1, which is in and is required for protein is consistent with the that proteins may be targeted through these S. R. M. 1986; PubMed Scopus Google Scholar). However, is as to the of proteins may be required for the of calpains (1Carillo S. Pariat M. Steff A.M. Roux P. Etienne-Julan M. Lorca T. Piechaczyk M. Oncogene. 1994; 9: 1679-1689PubMed Google M. J. E. J. Biol. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar, V. S. H. M. J. Biol. 1994; PubMed Scopus Google Scholar). of cleavage among the substrates of calpains is also of to YY1 may be differentially targeted by the proteasome and various YY1 previously (24Kalenik J.L. Chen D. Bradley M.E. Chen S.J. Lee T.C. Nucleic Acids Res. 1997; 25: 843-849Crossref PubMed Scopus (87) Google Scholar), we found that the zinc finger the both at the of YY1, was required for calpain the but not to calpain that the in play a role in calpain of YY1. findings that different structural are involved in and of evidence suggest that calpain II plays an important regulatory role in myogenic differentiation. The increase in calpain II activity with the of myoblast which is by cell membrane fusion and calcium influx K.B. Chung S.S. Kim O.M. Kang M.S. Ha D.B. Chung C.H. Biochim. Biophys. Acta. 1993; 1175: 243-249Crossref PubMed Scopus (72) Google Scholar). added calpain II was found to myoblast fusion J. N. D. A. M. S. P. A. J. Cell Biol. 1994; Google Scholar). calpain inhibitors also that of the endogenous calpain can myogenesis (14Kumar A. Shafiq S. Wadgaonkar R. Stracher A. Cell. Mol. Biol. 1992; 38: 687-691PubMed Google Scholar, K.B. J. Kang M.S. Ha D.B. Chung C.H. FEBS Lett. 1993; PubMed Scopus Google Scholar). Although of calpain II in muscle growth and differentiation have been postulated (6Goll D.E. Thompson V.F. Taylor R.G. Christiansen J.A. Biochimie (Paris). 1992; 74: 225-237Crossref PubMed Scopus (219) Google Scholar, Cell Res. 1986; PubMed Scopus Google Scholar), the mechanism myogenic function is unclear. The a regulatory mechanism for calpain II through proteolytic cleavage of the transcription factor YY1, which is capable of repressing myogenic transcription DNA-binding activity A. D. S. K. M. K. Mol. Cell. Biol. 1992; PubMed Scopus Google Scholar, T.C. Shi Y. Schwartz R.J. Proc. Natl. Acad. Sci. U. S. A. 1992; PubMed Scopus Google Scholar). in vivo evidence that YY1 can be proteolyzed by calpains is by the A23187 stabilization of YY1 protein by calpain inhibitor II in Consistent with this effect of this calcium ionophore was found to calpains J. E. J. Biol. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar) and myogenesis (17David J.D. See W.M. Higginbotham C. Dev. Biol. 1981; 82: 297-307Crossref PubMed Scopus (88) Google Scholar). The 26 S proteasome has been and found to be involved in the degradation of transcriptional as NF-κB (4Palombella V.J. Rando O.J. Goldberg A.L. Maniatis T. Cell. 1994; 78: 773-785Abstract Full Text PDF PubMed Scopus (1935) Google Scholar), M. D. Cell. 1994; 78: Full Text PDF PubMed Scopus Google Scholar), c-Fos (8Stancovski I. Gonen H. Orian A. Schwartz A.L. Ciechanover A. Mol. Cell. Biol. 1995; 15: 7106-7116Crossref PubMed Scopus (159) Google Scholar), p53 M. Huibregtse J.M. Howley P.M. Cell. Full Text PDF PubMed Scopus Google Scholar), and the repressor M. Ellison M.J. Chau V. Varshavsky A. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 4606-4610Crossref PubMed Scopus (208) Google Scholar). and L.J. Chen S.J. J.L. Bradley M.E. Lee T.C. Cell Res. 1996; PubMed Scopus Google Scholar) that YY1 turnover is mediated by calpains as well as the proteasome. We found that which is used as an inhibitor of the proteasome Goldberg A.L. J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar), is to YY1 in MG132 and calpain inhibitor II are both of and respectively. Thus, the of MG132 to calpains as shown may not be MG132 to be an inhibitor of both calpains and the it is that stabilization of YY1 in vivo can be observed under the both proteolytic pathways are This is consistent with the findings that the stabilization of YY1 in vivo by lactacystin can only be observed in myoblasts that proteasome L.J. Chen S.J. J.L. Bradley M.E. Lee T.C. Cell Res. 1996; PubMed Scopus Google Scholar). In cellular processes with calcium influx by A23187 calpain may a major proteolytic This may stabilization of YY1 by calpain inhibitors can only be observed in The of proteins are found to or in and S. R. M. 1986; PubMed Scopus Google Scholar). However, the mechanism that to rapid proteolysis remains unclear. The structural or of transcription factors that to be rapidly in vivo to be further We present evidence that calpains and the proteasome different structural elements for substrate finding the that not substrate to calpain proteolysis M. J. E. J. Biol. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar). targeting by the proteasome on the other involves the of These targeting mechanisms of calpains and the proteasome presumably provide elements of for cellular regulation and for a developmental control the pathway is well known for we that the proteasome and calpains may differentially the stability of a protein under different growth and as by the of A23187 and serum YY1, the stability of c-Fos and p53 has previously been found to be by calpains and the proteasome (8Stancovski I. Gonen H. Orian A. Schwartz A.L. Ciechanover A. Mol. Cell. Biol. 1995; 15: 7106-7116Crossref PubMed Scopus (159) Google Scholar, 9Hirai S. Kawasaki H. Yaniv M. Suzuki K. FEBS Lett. 1991; 287: 57-61Crossref PubMed Scopus (150) Google Scholar, 10Maki C.G. Huibregtse J.M. Howley P.M. Cancer Res. 1996; 56: 2649-2654PubMed Google Scholar, 11Kubbutat M.H.G. Vousden K.H. Mol. Cell. Biol. 1997; 17: 460-468Crossref PubMed Scopus (280) Google Scholar). from the of various proteases in the level of transcription the DNA-binding factor has been found to a protease activity A. H. 1995; PubMed Scopus Google Scholar). Thus, targeting proteases to a transcriptional may represent a in gene We Cecile Pickart for MG132 and
Walowitz et al. (Sun,) studied this question.