DMBT1 confers mucosal protection in vivo and a deletion variant is associated with Crohn's disease.

BACKGROUND & AIMS: Impaired mucosal defense plays an important role in the pathogenesis of Crohn's disease (CD), one of the main subtypes of inflammatory bowel disease (IBD). Deleted in malignant brain tumors 1 (DMBT1) is a secreted scavenger receptor cysteine-rich protein with predominant expression in the intestine and has been proposed to exert possible functions in regenerative processes and pathogen defense. Here, we aimed at analyzing the role of DMBT1 in IBD. METHODS: We studied DMBT1 expression in IBD and normal tissues by quantitative reverse transcription-polymerase chain reaction, immunohistochemistry, and mRNA in situ hybridization. Genetic polymorphisms within DMBT1 were analyzed in an Italian IBD case-control sample. Dmbt1(-/-) mice were generated, characterized, and analyzed for their susceptibility to dextran sulfate sodium-induced colitis. RESULTS: DMBT1 levels correlate with disease activity in inflamed IBD tissues. A highly significant fraction of the patients with IBD displayed up-regulation of DMBT1 specifically in the intestinal epithelial surface cells and Paneth cells. A deletion allele of DMBT1 with a reduced number of scavenger receptor cysteine-rich domain coding exons is associated with an increased risk of CD (P = .00056; odds ratio, 1.75) but not for ulcerative colitis. Dmbt1(-/-) mice display enhanced susceptibility to dextran sulfate sodium-induced colitis and elevated Tnf, Il6, and Nod2 expression levels during inflammation. CONCLUSIONS: DMBT1 may play a role in intestinal mucosal protection and prevention of inflammation. Impaired DMBT1 function may contribute to the pathogenesis of CD.

Primary cutaneous T-cell lymphomas show a deletion or translocation affecting NAV3, the human UNC-53 homologue.

Multicolor fluorescent in situ hybridization (FISH) was used to identify acquired chromosomal aberrations in 12 patients with mycosis fungoides or Sézary syndrome, the most common forms of primary cutaneous T-cell lymphoma (CTCL). The most frequently affected chromosome was 12, which showed clonal deletions or translocations with a break point in 12q21 or 12q22 in five of seven consecutive Sézary syndrome patients and a clonal monosomy in the sixth patient. The break point of a balanced translocation t(12;18)(q21;q21.2), mapped in the minimal common region of two deletions, fine mapped to 12q2. By locus-specific FISH, the translocation disrupted one gene, NAV3 (POMFIL1), a human homologue of unc-53 in Caenorhabditis elegans. A missense mutation in the remaining NAV3 allele was found in one of six cases with a deletion or translocation. With locus-specific FISH, NAV3 deletions were found in the skin lesions of four of eight (50%) patients with early mycosis fungoides (stages IA-IIA) and in the skin or lymph node of 11 of 13 (85%) patients with advanced mycosis fungoides or Sézary syndrome. Preliminary functional studies with lentiviral small interfering RNA-based NAV3 silencing in Jurkat cells and in primary lymphocytes showed enhanced interleukin 2 expression (but not CD25 expression). Thus, NAV3 may contribute to the growth, differentiation, and apoptosis of CTCL cells as well as to the skewing from Th1-type to Th2-type phenotype during disease progression. NAV3, a novel putative haploinsufficient tumor suppressor gene, is disrupted in most cases of the commonest types of CTCL and may thus provide a new diagnostic tool.

CRP-ductin, the mouse homologue of gp-340/deleted in malignant brain tumors 1 (DMBT1), binds gram-positive and gram-negative bacteria and interacts with lung surfactant protein D.

CRP-ductin is a protein expressed mainly by mucosal epithelial cells in the mouse. Sequence homologies indicate that CRP-ductin is the mouse homologue of human gp-340, a glycoprotein that agglutinates microorganisms and binds the lung mucosal collectin surfactant protein-D (SP-D). Here we report that purified CRP-ductin binds human SP-D in a calcium-dependent manner and that the binding is not inhibited by maltose. The same properties have previously been observed for gp-340 binding of SP-D. CRP-ductin also showed calcium-dependent binding to both gram-positive and -negative bacteria. A polyclonal antibody raised against gp-340 reacted specifically with CRP-ductin in Western blots. Immunoreactivity to CRP-ductin was found in the exocrine pancreas, in epithelial cells throughout the gastrointestinal tract and in the parotid ducts. A panel of RNA preparations from mouse tissues was screened for CRP-ductin and SP-D expression by reverse transcription-PCR. The pancreas was the main site of synthesis of CRP-ductin, but transcripts were also readily amplified from salivary gland, the gastrointestinal tract, liver, testis, uterus and lung. Lung was the main site of synthesis of SP-D, but transcripts were also amplified from uterus, salivary gland, thymus, thyroid gland, pancreas and testis. We conclude that CRP-ductin is the mouse homologue of human gp-340 and that its capacity to bind SP-D as well as gram-negative and gram-positive bacteria suggests a role in mucosal immune defense.

Frequent downregulation of DMBT1 and galectin-3 in epithelial skin cancer.

DMBT1 and galectin-3 are potential interacting proteins with presumably complex roles in tumorigenesis. While at present a variety of mechanisms are discussed for DMBT1 and its participation in cancer, galectin-3 is commonly known to exert tumor-promoting effects. However, in vitro studies in a rodent system have suggested that DMBT1/galectin-3 interaction in the ECM triggers epithelial differentiation, which would point to tumor-suppressive properties. To improve the understanding of DMBT1/galectin-3 action in cancer, we carried out studies in skin cancer of different origins. Mutational analyses of DMBT1 identified a missense mutation in 1 of 13 melanoma cell lines. It led to an exchange of an evolutionary conserved proline residue for serine and located within the second CUB domain of DMBT1. Immunohistochemical analyses demonstrated absence of DMBT1/galectin-3 expression from melanocytes but induction of DMBT1 expression in 1 of 8 nevi and 1 of 11 melanomas and of galectin-3 expression in 3 of 8 nevi and 4 of 8 melanomas. These data suggest that DMBT1 and galectin-3 are unlikely to act as classical tumor suppressors in melanomas. DMBT1 and galectin-3 appear to be secreted to the ECM by epithelial cells within the epidermis and the hair follicle. Compared to the flanking normal epidermis, skin tumors of epithelial origin frequently displayed downregulation of DMBT1 (18 of 19 cases) and galectin-3 (12 of 12 cases). Thus, loss of DMBT1/galectin-3 expression may play a role in the genesis of epithelial skin cancer. This would support the view that galectin-3 can exert tumor-suppressive effects in certain scenarios, and DMBT1/galectin-3-mediated differentiation represents a candidate mechanism for this effect.

An integrative model on the role of DMBT1 in epithelial cancer.

The gene, deleted in malignant brain tumors 1 (DMBT1), has been proposed to play a role in brain and epithelial cancer, but shows unusual features for a classical tumor suppressor gene. We have proposed that its presumptive dual function in protection and differentiation is of importance to understand its role in cancer. To gain insights into its role in tumorigenesis, we conducted a comprehensive study on DMBT1 mutations, expression and location. Twenty-one out of 44 tumors showed variable numbers of tandem repeats (VNTRs) due to genetic polymorphism of DMBT1, whereas 11 out of 44 tumors displayed presumable mutations. However, none of the alterations would be predicted to lead to a complete inactivation of the gene. DMBT1 is mucin-like and shows tissue-specific expression and secretion, pointing to a function in the protection of monolayered epithelia and to an additional function in the differentiation of multilayered epithelia. The expression patterns in carcinomas arising from the respective structures support this view. Accepting this functional dualism gives rise to an initial model on the role of DMBT1 in epithelial cancer.

The SRCR/SID region of DMBT1 defines a complex multi-allele system representing the major basis for its variability in cancer.

Deleted in malignant brain tumors 1 (DMBT1) at 10q25.3-q26.1 has been proposed as a candidate tumor-suppressor gene for brain and epithelial cancer. DMBT1 encodes a multifunctional mucin-like protein presumably involved in epithelial differentiation and protection. The gene consists of highly homologous and repeating exon and intron sequences. This specifically applies to the region coding for the repetitive scavenger receptor cysteine-rich (SRCR) domains and SRCR-interspersed domains (SIDs) that constitutes the major part of the gene. This particular structure may previously have interfered with the delineation of DMBT1 alterations in cancer. Uncovering these, however, is of mechanistic importance. By a combined approach, we conducted a detailed mutational analysis, starting from a panel of 51 tumors, including 46 tumor cell lines and five primary tumors. Alterations in the repetitive region were present in 22/31 (71%) tumors that were investigated in detail. Six tumors showed presumably de novo mutations, among these three with point mutations in combination with a loss of heterozygosity. However, none of the alterations unambiguously would be predicted to lead to an inactivation of DMBT1. We define seven distinct DMBT1 alleles based on variable numbers of tandem repeats (VNTRs). At least 11 tumors exclusively harbored these VNTRs. The data suggest that the SRCR/SID region defines a complex multi-allele system that has escaped previous analyses and that represents the major basis for the variability of DMBT1 in cancer. DMBT1 thus compares to mucins rather than to conventional tumor suppressors.

Sequential changes of the DMBT1 expression and location in normal lung tissue and lung carcinomas.

Deleted in Malignant Brain Tumors 1 (DMBT1) at chromosome region 10q25.3-q26.1 has been proposed as a candidate tumor-suppressor gene for brain, digestive tract, and lung cancer. Recent studies on its expression in lung cancer have led to divergent results and have raised a controversial discussion. Moreover, DMBT1 has been implicated with epithelial protection in the respiratory tract. We thus wondered how a loss of its expression could be related to carcinogenesis in the lung. To address these issues, we investigated the DMBT1 expression and location in the normal lung and lung cancer. By reverse-transcription PCR, a down-regulation of the DMBT1 expression in lung cancer cell lines is commonly detected. Immunohistochemical studies in situ demonstrate that there are also low steady-state levels of DMBT1 in the normal respiratory epithelium. However, an up-regulation takes place in the tumor-flanking epithelium and upon respiratory inflammation. Lung carcinomas show increased DMBT1 expression compared to that of undiseased lung tissue, but decreased DMBT1 levels compared to that of tumor-flanking and inflammatory tissue. A switch from a lumenal secretion to a secretion to the extracellular matrix takes place during lung carcinogenesis. Our data may resolve the controversial discussion on its expression in lung carcinomas. We hypothesize that the changes of the DMBT1 expression and location do reflect a time course that may point to possible mechanisms for its role in epithelial cancer.