Biomedical subjects

R L Eckert

Publications and source records attributed to R L Eckert.

At least 37 records · Page 2Linked to original sources

PubMed1998

The distal regulatory region of the human involucrin promoter is required for expression in epidermis.

Human involucrin (hINV) is a precursor of the keratinocyte cornified envelope that is specifically expressed in the suprabasal layers of stratifying squamous epithelia. The promoter distal (DRR) and proximal regulatory regions (PRR) are required for optimal in vitro expression (Welter, J. F., Crish, J. F., Agarwal, C., and Eckert, R. L. (1995) J. Biol. Chem. 270, 12614-12622; and Banks, E. B., Crish, J. F., Welter, J. F., and Eckert, R. L. (1998) Biochem. J. 331, 61-68). We now present the complete sequence of these regions and evaluate their ability to drive in vivo transcription. Transgenes containing 5000 or 2473 base pairs of upstream regulatory region drive tissue- and differentiation-appropriate expression in stratifying surface epithelia. In contrast, transgenes containing 1953, 1333, 986, or 41 base pairs of upstream regulatory region are not expressed in surface epithelia, indicating that loss of the DRR (nucleotides -2474/-1953) results in loss of expression. Fusing the isolated DRR region directly to the hINV minimal promoter restores surface epithelial expression. Sequences downstream of the transcribed gene are not required for appropriate expression. The -1953/-41 segment influences the pattern of differentiation-dependent expression. The -986/-41 region, which includes the PRR, drives expression in internal epithelia.

Animals↗

PubMed1998

Transcriptional regulation of the EGF receptor promoter by HPV16 and retinoic acid in human ectocervical epithelial cells.

We have previously demonstrated that human papillomavirus 16 (HPV16)-immortalized human ectocervical epithelial cells and cells derived from tumors which express HPV16 oncogenes express high levels of epidermal growth factor receptor (EGFR) compared to normal cervical cells. We have also shown that proliferation of these cells is inhibited by retinoid treatment. We have hypothesized that the retinoid inhibition of cell proliferation may be due to the retinoid-dependent reduction in EGFR level. In this study we examine the regulation of EGFR expression in cervical cells with emphasis on two aspects: (1) the mechanism of retinoid-dependent suppression of EGFR levels in HPV16-positive cells and (2) the mechanism of EGFR upregulation by HPV16. EGFR levels were found to be elevated 5-, 3. 7-, and 1.25-fold in the HPV16-immortalized ECE16-1, ECE16-D1, and ECE16-D2 cells, respectively, compared to normal cervical cells. Treatment of ECE16-1 and ECE16-D1 cells with retinoic acid suppresses proliferation, EGFR level, EGFR mRNA level, and EGFR promoter activity. The reduction in EGFR promoter activity appears to account for the reduction in EGFR protein and mRNA levels. In contrast, retinoic acid does not affect cell growth or EGFR level in ECE16-D2 cells or normal cervical cells. To study the mechanisms regulating EGFR expression in HPV16-positive cells, normal ECE cells were cotransfected with an EGFR promoter reporter plasmid and an expression plasmid encoding the HPV16 E6/E7 open reading frames. In the presence of E6/E7, EGFR promoter activity was increased by 2- to 3-fold, suggesting that the E6/E7 proteins are directly or indirectly responsible for the increased EGFR level and that the EGFR promoter contains the DNA elements necessary to mediate this response. Nevertheless expression of E6/E7 proteins did not confer retinoic acid regulation, as EGFR promoter activity remained elevated in normal cells cotransfected with pHPVE6/E7 and treated with retinoic acid. These results suggest that human papillomavirus and retinoic acid regulate EGFR levels by independent effects on the EGFR promoter.

Cell Division↗

PubMed1998

Regulation of human involucrin promoter activity by a protein kinase C, Ras, MEKK1, MEK3, p38/RK, AP1 signal transduction pathway.

Involucrin is a marker of keratinocyte terminal differentiation. Our previous studies show that involucrin mRNA levels are increased by the keratinocyte differentiating agent, 12-O-tetradecanoylphorbol-13-acetate (TPA) (Welter, J. F., Crish, J. F., Agarwal, C., and Eckert, R. L. (1995) J. Biol. Chem. 270, 12614-12622). We now study the signaling cascade responsible for this regulation. Protein kinase C and tyrosine kinase inhibitors inhibit both the TPA-dependent mRNA increase and the TPA-dependent increase in hINV promoter activity. The relevant response element is located within the promoter proximal regulatory region and includes an AP1 site, AP1-1. Co-transfection of the hINV promoter with dominant negative forms of Ras, MEKK1, MEK1, MEK7, MEK3, p38/RK, and c-Jun inhibit the TPA-dependent increase. Wild type MEKK1 enhances promoter activity and the activity can be inhibited by dominant negative MEKK1, MEK1, MEK7, MEK3, p38/RK, and c-Jun. In contrast, wild type Raf-1, ERK1, ERK2, MEK4, or JNK1 produced no change in activity and the dominant negative forms of these kinases failed to suppress TPA-dependent transcription. Treatment with an S6 kinase (S6K) inhibitor, or transfection with constitutively active S6K produced relatively minor changes in promoter activity, ruling out a regulatory role for S6K. These results suggest that activation of involucrin transcription involves a pathway that includes protein kinase C, Ras, MEKK1, MEK3, and p38/RK. Additional pathways that transfer MEKK1 activation via MEK1 and MEK7 also may function, but the downstream targets of these kinases need to be identified. AP1 transcription factors appear to be the ultimate target of this regulation.

Base Sequence↗

PubMed1998

Characterization of human involucrin promoter distal regulatory region transcriptional activator elements-a role for Sp1 and AP1 binding sites.

Human involucrin (hINV) is an important precursor of the keratinocyte cornified envelope that is specifically expressed in the suprabasal layers of stratifying epithelia. Previous truncation and mutagenesis experiments have shown that an activator protein 1 (Ap1) site, AP1-5, located 2100bp upstream of the transcription start site, is required for optimal promoter activity. These previous studies suggest that AP1-5 is part of a distal regulatory region spanning nucleotides -2473 to -2088. In the present report, we study the distal regulatory region (DRR), which surrounds AP1-5. Our studies show that this region contains weak and strong activator elements spanning nucleotides -2473/-2216 and -2140/-2088, respectively. The strong activator element contains AP1-5 and an adjacent specificity protein 1 (Sp1) site. The AP1-5 site is absolutely required for DRR activity, as its mutation reduces transcription to basal levels. Mutagenesis studies of the AP1-5 and Sp1 sites in the presence or absence of the weak activator element indicate that the Sp1 site and the weak activator element synergistically activate the AP1-5 site-dependent transcription. The cooperation between the Sp1 and AP1-5 sites is also observed in the context of the full-length promoter. Gel mobility shift and supershift studies show that Sp1, but not Sp2, Sp3 or Sp4 binds to the Sp1 site. When the Sp1 site is mutated or the distance between the AP1-5 and Sp1 site is increased, the binding of AP1 factors to AP1-5 is markedly reduced. Surprisingly, gel shift studies suggest that activation does not require the formation of a stable AP1/Sp1/DNA ternary complex. These studies suggest that the AP1-5 site is absolutely required for transcriptional activation, that the weak activator element and Sp1 sites serve to enhance this activation, and that the Sp1 site is required for optimal AP1 factor binding at the AP1-5 site.

Base Sequence↗

PubMed1998

Identification of transglutaminase-reactive residues in S100A11.

The recent finding that S100A11 is a component of the keratinocyte cornified envelope (CE) (Robinson, N. A., Lapic, S., Welter, J. F., and Eckert, R. L. (1997) J. Biol. Chem. 272, 12035-12046) suggests that S100A11 is a transglutaminase (TG) substrate. In the present study we show that S100A11 forms multimers when cultured keratinocytes are challenged by increased levels of intracellular calcium and that multimer formation is inhibited by the TG inhibitor, cystamine. These S100A11 multimers appear to be incorporated into the CE, as immunoreactive S100A11 is detected in purified envelopes prepared from cultured cells and from foreskin epidermis. To study S100A11 as a transglutaminase substrate, recombinant human S100A11 (rhS100A11) was used in a cell-free cross-linking system. [14C]Putrescine, a primary amine, labels rhS100A11 in a TG-dependent manner. Trypsin digestion of [14C]putrescine-labeled rhS100A11 releases one radiolabeled peptide, Ala98-Lys103. The glutamine residue in this segment, Gln102, is the site of radiolabel incorporation indicating that Gln102 functions as an amine acceptor. The ability of S100A11 to form multimers indicates that it also has a reactive lysine residue that functions as an amine donor. To identify the reactive residue, we compared the high pressure liquid chromatography profile of trypsin-digested rhS100A11 monomer to that of cross-linked rhS100A11. A unique cross-linked peptide was purified and identified as Met-Ala-Lys3-Ilu-Ser-Ser-Pro-Thr-Glu-Thr-Glu-Arg cross-linked via an Lys3-Gln102 isopeptide bond to Ala-Val-Pro-Ser-Gln102-Lys. These studies show that S100A11 is post-translationally modified by transglutaminase, that it can be cross-linked to form multimers, that it is present in CEs from cultured keratinocytes and in vivo epidermis, and that Lys3 and Gln102 are specific sites of cross-link formation.

Cell Compartmentation↗

PubMed1998

Retinoids modulate P2U purinergic receptor-mediated changes in transcervical paracellular permeability.

In human cervical cells, extracellular ATP induces an acute decrease in the resistance of the lateral intercellular space, the phase I response, followed by a delayed increase in tight junctional resistance, the phase II response. These responses depend on vitamin A because incubation of cells in retinoid-free medium (RFM) abolished both responses. Treatment with retinoic acid restored the phase I response in full, but the amplitude of the phase II response was restored only partly. Shorter incubations and lower concentrations of retinoic acid [half-maximal effective concentrations (K 1/2) = 0.1 microM] were required for restoring the phase I response than were required for reversing the phase II response (K 1/2 = 1 microM). The phase I response could be restored by ligands that bind to either retinoic acid receptors (RARs) or retinoid X receptors, but only RAR agonists had an effect on phase II response. RFM had no effect on decreases in resistance induced by ionomycin, but it attenuated phase II-like increases in resistance induced by KCl or by 1,2-dioctanoyl-sn-diglycerol (diC8). Actinomycin D blocked phase II response but not phase I response or the responses to ionomycin, KCl, or diC8. These results suggest that retinoids act on cervical cells via distinct retinoid receptor mechanisms and modulate phase I and phase II changes in resistance by regulating distinct signal mechanisms.

Adenosine Triphosphate↗

PubMed1998

In vitro cross-linking of recombinant human involucrin.

Human involucrin (hINV) is a constituent of the scaffolding of the cornified envelope. In the present study, we describe an in vitro model system to study the role of hINV in scaffold formation. We characterize the in vitro cross-linking of full-length (585 amino acid) recombinant hINV, rhINV(1-585). When reacted with detergent-solubilized, particulate transglutaminase type 1 (TG1) or partially purified type 2 transglutaminase (TG2), rhINV(1-585) functions as a TG substrate in a calcium-dependent manner. When the reaction is supplemented with 14C-putrescine tracer, the radiolabeled cosubstrate is incorporated into a high-molecular-weight product in a calcium-, rhINV(1-585)- and time-dependent manner. 35S-rhINV(1-585) is also cross-linked to form a high-molecular-weight product. These results suggest that rhINV(1-585) is extensively multimerized. Products having a molecular weight smaller than authentic rhINV(1-585) are also formed, providing evidence for intramolecular cross-link formation. Transmission electron microscopy of cross-linked product reveals immunoreactive large-molecular-weight loop-string-loop and branched structures. Our studies (1) show that rhINV(1-585) is a substrate for both TG1 and TG2, (2) indicate that rhINV(1-585) can be cross-linked to form macromolecular products having distinct structural features, (3) demonstrate that rhINV(1-585) forms intramolecular cross-links when hINV concentration is limiting and (4) establish that hINV possesses reactive Gln and Lys residues.

Bacteria↗

PubMed1997

S100A11, S100A10, annexin I, desmosomal proteins, small proline-rich proteins, plasminogen activator inhibitor-2, and involucrin are components of the cornified envelope of cultured human epidermal keratinocytes.

The cornified envelope (CE) is an insoluble sheath of epsilon-(gamma-glutamyl)lysine cross-linked protein, which is deposited beneath the plasma membrane during keratinocyte terminal differentiation. We have probed the structure of the CE by proteolytic cleavage of purified CE fragments isolated from CEs formed spontaneously in cell culture. CNBr digestion, followed by trypsin and then proteinase K treatment released 25%, 42%, and 18%, respectively, of the CE protein. Purification and sequencing of released peptides has identified two novel CE precursors, S100A11 (S100C, calgizzarin) and S100A10 (calpactin light chain). We also sequenced peptides derived from annexin I and plasminogen activator inhibitor 2, two putative envelope precursors, as well as portions of the well established CE precursor proteins SPR1A, SPR1B, and involucrin. Many desmosomal components were identified (desmoglein 3, desmocolin A/B, desmoplakin I, plakoglobin, and plakophilin), indicating that desmosomes become cross-linked into the CE. Fragments derived from envoplakin, the recently sequenced 210-kDa membranous CE precursor protein, which also appears to be a desmosomal component, were also identified. Analysis of the pattern of peptide release following the sequential digestion indicates that S100A11 is anchored to the envelope via Gln102 and/or Lys103 at the carboxyl terminus and at Lys3, Lys23, and/or Gln22 in the amino terminus. A similar type of analysis indicates that small proline-rich proteins 1A and 1B (SPR1A and SPR1B) become cross-linked at the amino terminus (residues 1-23) and the carboxyl terminus (residues 86-89). No loricrin, cystatin A, or elafin peptides were detected.

Amino Acid Sequence↗

PubMed1997

The epidermis: genes on - genes off.

The epidermal keratinocyte stem cell is distinguished by a relatively undifferentiated phenotype and an ability to proliferate. As part of a carefully orchestrated process, the offspring of these stem cells lose the ability to proliferate and begin a process of morphologic and biochemical transformation that results in their conversion into corneocytes. This process requires the coordinated expression of a host of cellular genes. The mechanisms responsible for regulation of these genes is an area of intense interest. In keratinocytes, as in other cell types, the expression of most genes is regulated at the transcriptional level by a class of proteins called transcription factors. Transcription factors are nuclear proteins that regulate transcription by mediating the final steps in the relay of information from the cell surface to the nucleus and the gene. These factors bind to specific DNA sequence elements located within the target gene. In this brief review we summarize evidence implicating activator protein 1 (AP1), AP2, Sp1, POU domain, CCAAT enhancer binding protein, and several other transcription factors as regulators of expression of keratinocyte genes.

Animals↗

PubMed1997

Retinoids regulate tight junctional resistance of cultured human cervical cells.

The objective of the study was to determine the effect of retinoids on paracellular resistance across the cervical epithelium and the mechanisms involved. The experimental model was cultures of human CaSki cells on filters, which retain phenotypic characteristics of the endocervical epithelium. End points for paracellular resistance were measurements of transepithelial electrical resistance and fluxes of pyranine (a trisulfonic acid that traverses the epithelium via the intercellular space). Paracellular resistance was significantly increased in cells grown in retinoid-free medium; the effect could be blocked and reversed with all-trans-retinoic acid (tRA) and with agonists of RAR and RXR receptors but only partially with retinol. The effect of tRA was dose dependent and saturable, with a 50% effective concentration of 0.8 nM. The increases in paracellular resistance induced by vitamin A deficiency required longer incubation in retinoid-free medium than decreases in resistance induced by retinoic acid. tRA had only a minimal effect on paracellular resistance in cells maintained in regular medium. Retinoid-free medium increased and tRA decreased the relative cation mobility across CaSki cultures. Also the effects of tRA were nonadditive to those of cytochalasin D (which decreases tight junctional resistance) and additive to those of ionomycin (which decreases the resistance of the lateral intercellular space), suggesting that tRA modulates tight junctional resistance. It is concluded that vitamin A determines the degree of paracellular resistance across cervical cells by a mechanism that involves modulation of tight junctional resistance.

Cell Line↗

PubMed1997

The epidermal keratinocyte as a model for the study of gene regulation and cell differentiation.

The epidermis is a dynamic, continually renewing structure that provides the organism with a life-sustaining interface with the environment. The major cell type of the epidermis, the epidermal keratinocyte, undergoes a complex and carefully choreographed program of differentiation. Aberrations in this process result in the genesis of a variety of debilitating and life-threatening diseases. In the present paper, we discuss the keratinocyte differentiation program and the exogenous agents that regulate differentiation. We describe the marker genes that have been utilized to study the process of gene regulation in epidermis. We describe the keratin proteins and studies that have identified keratin mutations that cause epidermal disease. We present recent information on regulation of keratinocyte gene expression and attempt to summarize current knowledge on the role of transcription factors in this process. We also discuss the process of cornified envelope assembly and the structure and function of the proteins that are thought to be precursors of this structure.

Animals↗

PubMed1996

Regulation of human involucrin promoter activity by POU domain proteins.

POU domain transcription factors are expressed in the epidermis and are thought to be important regulators of keratinocyte gene expression. In the present article we demonstrate that POU transcription factors suppress transcription of the human involucrin (hINV) promoter. Cotransfection of pINV-2473, a construct containing 2473 base pairs of hINV upstream sequence linked to luciferase, with POU homeodomain transcription factors Oct1, Oct2, Brn4, SCIP, Skn1a or Skn1i, results in a strong suppression of basal promoter activity. The hINV upstream region includes a consensus POU transcription factor binding site, 5'-ATGCAAAT-3', centered around nucleotide -1277. Although this site interacts with POU factors, assays of promoter activity for a series of progressive 5' end truncations demonstrate that this site is not required for POU factor-dependent transcriptional suppression. Suppression is observed with the shortest truncation construct tested, pINV-41, suggesting that this inhibition may be mediated by effects on TATA box proteins. SCIP mutants that lack transactivation or DNA binding domains were shown to suppress transcription, suggesting that the DNA binding and transactivation domains are not required for suppression. Moreover, cotransfection of the pINV-2473 with pKSM13(+)OCT, which contains a single consensus OCT binding site, results in an increase in basal promoter activity, suggesting that endogenous POU factors suppress hINV promoter activity. In addition to inhibiting basal transcription, POU transcription factors also suppress phorbol ester-stimulated hINV promoter activity. These studies suggest that suppression of hINV promoter activity does not require the amino-terminal segment of the POU factor or direct POU factor interaction with DNA and suggest that the suppression may be via indirect interaction with other proteins in the vicinity of the TATA box. Thus, involucrin joins the ranks of a small set of genes that are regulated by POU factors in an octamer binding site-independent manner.

Base Sequence↗

PubMed1996

AGN193109 is a highly effective antagonist of retinoid action in human ectocervical epithelial cells.

Retinoids are important physiological agents that regulate epithelial cell differentiation and proliferation. The importance of these agents in regulating growth, development, and differentiation has led to a search for new retinoid agonists and antagonists. In the present manuscript we show that AGN193109, a retinoid analog, is an efficient antagonist of retinoid action in human cervical epithelial cells. Treatment of ECE16-1 cells with natural or synthetic retinoids reduces cytokeratin K5, K6, K14, K16, and K17 levels, increases cytokeratin K7, K8, and K19 levels, increases retinoic acid receptor-beta (RAR beta) mRNA levels, suppresses proliferation, and alters cell morphology. Co-treatment with AGN193109 prevents these responses. Half-maximal and maximal antagonism is observed at a molar ratio of AGN193109: retinoid agonist of 1:1 and 10:1, respectively. When administered alone AGN193109 has no agonist activity. Thus, AGN193109, which binds to RAR alpha, RAR beta, and RAR gamma with Kd values = 2,2, and 3 nm, respectively, but is unable to bind to the retinoid X receptors, is a highly active antagonist of retinoid action in ECE16-1 cells.

Cell Division↗

PubMed1996

Regulation of human dermal papilla cell production of insulin-like growth factor binding protein-3 by retinoic acid, glucocorticoids, and insulin-like growth factor-1.

Insulin-like growth factor-1 (IGF1) has been reported to stimulate hair elongation and to facilitate maintenance of the hair follicle in anagen phase. However, little is known about IGF1 signaling in the hair follicle. In this study we investigate the effects of IGF1, glucocorticoids, and retinoids on dermal papilla (DP) cell production of insulin-like growth factor binding proteins (IGFBPs). IGFBPs comprise a family of IGF binding proteins that are produced and released by most cell types. They bind to IGFs to either enhance or inhibit IGF activity. In the present report we identify IGFBP-3 as being produced and released by cultured human dermal papilla (DP) cells. IGFBP-3 levels are increased fivefold by retinoic acid, eightfold by dexamethasone, and tenfold by IGF1. DP cells are known to produce IGF1, and so the observed stimulation of DP cell IGFBP-3 production by IGF1 is consistent with the idea that DP cells possess the IGF transmembrane receptor kinase and are autoregulated by IGFs. The level of another IGFBP, tentatively identified as IGFBP-2, is, in contrast, not regulated by these agents. IGFBP-3 has been shown to inhibit the activity of IGFs in a variety of systems. Our results are consistent with a model in which retinoids and glucocorticoids inhibit IGF action on DP cells and surrounding matrix cells by stimulating increased DP cell production of IGFBP-3. The IGFBP-3, in turn, forms a complex with free IGF1 to reduce the concentration of IGF1 available to stimulate hair elongation and maintenance of anagen phase.

Cells, Cultured↗

PubMed1996

Transcription factor regulation of epidermal keratinocyte gene expression.

The epidermis is a tissue that undergoes a very complex and tightly controlled differentiation program. The elaboration of this program is generally flawless, resulting in the production of an effective protective barrier for the organism. Many of the genes expressed during keratinocyte differentiation are expressed in a coordinate manner; this suggests that common regulatory models may emerge. The simplest model envisions a 'common regulatory element' that is possessed by all genes that are regulated together (e.g., involucrin and transglutaminase type 1). Studies to date, however, have not identified any such elements and, if anything, the available studies suggest that appropriate expression of each gene is achieved using sometime subtly and sometime grossly different mechanisms. Recent studies indicate that a variety of transcription factors (AP1, AP2, POU domain. Sp1, STAT factors) are expressed in the epidermis and, in many cases, multiple members of several families are present (e.g., AP1 and POU domain factors). The simultaneous expression of multiple members of a single transcription factor family provides numerous opportunities for complex regulation. Some studies suggest that specific members of these families interact with specific keratinocyte genes. The best studied of these families in epidermis is the AP1 family of factors. All of the known AP1 factors are expressed in epidermis [52] and each is expressed in a specific spatial pattern that suggests the potential to regulate multiple genes. It will be important to determine the role of each of these members in regulating keratinocyte gene expression. Finally, information is beginning to emerge regarding signal transduction in keratinocytes. Some of the early events in signal transduction have been identified (e.g., PLC and PKC activation, etc.) and some of the molecular targets of these pathways (e.g., AP1 transcription factors) are beginning to be identified. Eventually we can expect to elucidation of all of the steps between the interaction of the stimulating agent with its receptor and the activation of target gene expression.

Animals↗

PubMed1996

Involucrin is a covalently crosslinked constituent of highly purified epidermal corneocytes: evidence for a common pattern of involucrin crosslinking in vivo and in vitro.

Involucrin (hINV) is an important structural component of the keratinocyte cornified envelope that is expressed early in the keratinocyte differentiation process and is thought to be a component of the initial envelope scaffolding. We have previously shown that cyanogen bromide (CNBr) cleavage of cornified envelopes isolated from cultured foreskin keratinocytes releases several discrete involucrin-immunoreactive peptides. In this study, we compare the pattern of release of immunoreactive hINV fragments from envelopes prepared from human breast skin and foreskin, and from spontaneous and induced envelopes prepared from cultured keratinocytes. We also identify one of the released products. Envelopes prepared from human breast skin or foreskin, or spontaneous or induced envelopes prepared from cultured cells differ significantly in structure. The envelopes isolated from epidermis appear to be structurally complete, whereas spontaneous envelopes appear less complete and the induced envelopes appear to be the least complete. In spite of these structural differences, CNBr cleavage releases an identical quartet of hINV-immunoreactive peptides migrating between 68 and 81 kDa from each preparation. Immunoblots indicate that the quantity of hINV-immunoreactive material released per microg of envelope protein is as follows: induced > spontaneous > foreskin > breast skin. The fastest migrating peptide (68 kDa) comigrates with a peptide that is released after CNBr cleavage of bacterially produced-recombinant hINV. Amino-terminal amino acid sequencing of this peptide from recombinant hINV and from the cornified envelopes yields the sequence G-Q-L-K-H-L-E-Q-Q-E-G-Q-P-K-H. These results suggest that this fragment is the 275-amino acid segment of hINV beginning at G311 and extending to K585, and that this peptide is not crosslinked to another protein. These results indicate that a population of the envelope-associated hINV present in cultured and in vivo keratinocytes is crosslinked in the amino-terminal half. It is possible that this species represents an early intermediate in the involucrin crosslinking process.

Amino Acid Sequence↗

PubMed1996

Immortalization of epithelial cells.

The methodologies for isolating cell lines have become very powerful, particularly in terms of retaining differentiated features of the parent cells. Cell lines can be developed from primary or early passage cells as well as from transgenic animals that carry an immortalizing gene. Cell lines from epithelia have been selected for their polar orientation, tight junction formation, and expression of differentiated markers or functions. These cell lines provide useful models for studying cell biology of specific tissues, tumorigenicity, genetic abnormalities, or to help screen for effective methods of gene therapy.

Animals↗

PubMed1996

Epidermal keratinocytes - genes and their regulation.

In recent years, the human epidermal keratinocyte has been extensively studied. These studies have shown that epidermal growth factor (EGF), transforming growth factor-beta (TGFbeta), retinoids, phorbol ester, vitamin D and other agents regulate keratinocyte proliferation, differentiation, apoptosis and gene expression. We review progress in understanding the mechanisms that regulate keratinocyte structural gene expression. In most cases little is known regarding the factors that regulate gene expression in response to a physiological agent. However, the available results suggest a role for a variety of transcription factors, including STAT factors, NFkappaB, octamer site (OCT) binding proteins and activator protein 1 (AP1) factors in regulating expression of these genes. Among these transcriptional regulators, AP1 appears to play a central role. We review the current literature regarding the regulation of involucrin, loricin, transglutaminase type 1 and cytokeratin gene expression. This survey indicates that the AP1 family of transcriptional regulators is implicated in the regulation of nearly all of these genes. We also discuss recent studies which describe the distribution of the AP1 factors, c-jun, junB, junD, Fra-1,Fra-2, c-fos and fosB, in epidermis.

Journal Article↗