Titin is a giant elastic protein in vertebrate striated muscles with an unprecedented molecular mass of 3–4 megadaltons. Single molecules of titin extend from the Z-line to the M-line. Here, we define the molecular layout of titin within the Z-line; the most NH2-terminal 30 kD of titin is located at the periphery of the Z-line at the border of the adjacent sarcomere, whereas the subsequent 60 kD of titin spans the entire width of the Z-line. In vitro binding studies reveal that mammalian titins have at least four potential binding sites for α-actinin within their Z-line spanning region. Titin filaments may specify Z-line width and internal structure by varying the length of their NH2-terminal overlap and number of α-actinin binding sites that serve to cross-link the titin and thin filaments. Furthermore, we demonstrate that the NH2-terminal titin Ig repeats Z1 and Z2 in the periphery of the Z-line bind to a novel 19-kD protein, referred to as titin-cap. Using dominant-negative approaches in cardiac myocytes, both the titin Z1-Z2 domains and titin-cap are shown to be required for the structural integrity of sarcomeres, suggesting that their interaction is critical in titin filament–regulated sarcomeric assembly.

REFERENCES 1 Tagami H, Iwatsuki K, Iwase Y et al. Subcorneal pustular dermatosis with vesiculo-buUous eruption: Demonstration of subcorneal IgA deposits and leukocyte chemotactic factor. Br J Dermatol 1983; 109: 581-7. 2 Hashimoto T, Inamoto N, Nakamura K et al. Intercellular IgA dermatosis with clinical features of subcorneal pustular dermatosis. Arch Dermatol 1987; 123: 1062-5. 3 Thivolet J, Schmitt D. Langerhans cells and T cell subset identification using monoclonal antibodies: Immunocytochemical methods in light and electron microscopy applied on human skin. In: Immunopathology of the Skin (Beutner EH, Chorzelski TP, Kumar V, eds) 3rd edn.. New York: John Wiley & Sons, Inc. 1987; 125-52. 4 Marsden JR, Millard LG. Pyoderma gangrenosum, subcorneal pustular dermatosis and IgA paraproteinaemia. BrJ Dermatol 1986; 114: 125-9. 5 Wallach D, Cottenot F, Pelbois G et al. Subcorneal pustular dermatosis and monoclonal IgA.

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The development of apoptosis resistance appears to be an important factor in colon carcinogenesis. To gain an understanding of the molecular pathways altered during the development of apoptosis resistance, we selected three cell lines for resistance to induction of apoptosis by deoxycholate, an important etiologic agent in colon cancer. We then evaluated gene expression levels for 825 proteins in these resistant lines, compared with a parallel control line not subject to selection. Eighty-two proteins were identified as either over-expressed or under-expressed in at least two of the resistant lines, compared with the control. Thirty-five of the 82 proteins (43%) proved to have a known role in apoptosis. Of these 35 proteins, 21 were over-expressed and 14 were under-expressed. Of those that were over-expressed 18 of 21 (86%) are anti-apoptotic in some circumstances, of those that were under-expressed 11 of 14 (79%) are pro-apoptotic in some circumstances. This finding suggests that apoptosis resistance during selection among cultured cells, and possibly in the colon during progression to cancer, may arise by constitutive over-expression of multiple anti-apoptotic proteins and under-expression of multiple pro-apoptotic proteins. The major functional groups in which altered expression levels were found are post-translational modification (19 proteins), cell structure (cytoskeleton, microtubule, actin, etc.) (17 proteins), regulatory processes (11 proteins) and DNA repair and cell cycle checkpoint mechanisms (10 proteins). Our findings, overall, bear on mechanisms by which apoptosis resistance arises during progression to colon cancer and suggest potential targets for cancer treatment. In addition, assays of normal-appearing mucosa of colon cancer patients, for over- or under-expression of genes found to be altered in our resistant cell lines, may allow identification of early biomarkers of colon cancer risk.

Any pathogenetic mechanism proposed for erythema multiforme (EM) must account for the prominent mononuclear cell infiltrate in the skin lesions. The purpose of this study was to characterize immunopathologically, with monoclonal antibodies to human leukocyte antigens, the inflammatory cells in early target lesions of recurrent herpes-associated EM. Cryostat sections of snap-frozen skin biopsies were studied by the avidin-biotin immunoperoxidase technique with use of the following monoclonal antibodies: anti-HLA-DR, anti-Leu M5, anti-Leu 4 + 5b, anti-Leu 3a + 3b, anti-Leu 2a, anti-Leu 14, and anti-Leu 6. The dermal mononuclear inflammatory infiltrate in the EM biopsies consisted of monocyte-macrophages and T-lymphocytes, with both helper and suppressor T cells present. Both the dermal inflammatory infiltrate and the overlying keratinocytes were strongly HLA-DR positive. No definite alteration of Langerhans cell number or distribution was noted. These findings are consistent with the characteristics seen in cell-mediated immune reactions in the skin and point to this as a likely immune mechanism for the tissue damage of EM.

Deoxycholate, a bile salt present at high levels in the colonic lumen of individuals on a high-fat diet, is a promoter of colon cancer. Deoxycholate also causes DNA damage. BRCA-1 functions in repair of DNA and in induction of apoptosis. We show that, when cultured cells of colonic origin are exposed to deoxycholate at different concentrations, BRCA-1 expression is induced at a low noncytotoxic concentration (10 microM) but is strongly inhibited at higher cytotoxic concentrations ( > or =100 microM). Indication of phosphorylation of BRCA-1 by deoxycholate (100 microM) at a lower dose was seen by Western blot analysis, whereas, at a higher dose, deoxycholate (200 and 300 microM) caused a complete loss of BRCA-1 expression. We show that BRCA-1 is substantially lower in colon adenocarcinomas from five patients compared with associated non-neoplastic colon tissue from the same patients, suggesting that the loss of BRCA-1 expression contributes to the malignant phenotype. In the non-neoplastic colon tissue, BRCA-1 was localized to the nongoblet cells. Our results imply that reduced expression of BRCA-1 may be associated with carcinoma of the colon.

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