ABSTRACT
The t(15;17) translocation, found in 95% of acute promyelocytic leukemia, encodes a promyelocytic leukemia (PML)-retinoic acid receptor alpha (RARalpha) fusion protein. Complete remission of acute promyelocytic leukemia can be obtained by treating patients with all-trans retinoic acid, and PML-RARalpha plays a major role in mediating retinoic acid effects in leukemia cells. A main model proposed for acute promyelocytic leukemia is that PML-RARalpha exerts its oncogenic effects by repressing the expression of retinoic acid-inducible genes critical to myeloid differentiation. By applying subtraction cloning to acute promyelocytic leukemia cells, we identified a retinoic acid-induced gene, PRAM-1 (PML-RARalpha target gene encoding an Adaptor Molecule-1), which encodes a novel adaptor protein sharing structural homologies with the SLAP-130/fyb adaptor. PRAM-1 is expressed and regulated during normal human myelopoiesis. In U937 myeloid precursor cells, PRAM-1 expression is inhibited by expression of PML-RARalpha in the absence of ligand and de novo superinduced by retinoic acid. PRAM-1 associates with other adaptors, SLP-76 and SKAP-55HOM, in myeloid cell lines and with protein tyrosine kinase lyn. By providing the first evidence that PML-RARalpha dysregulates expression of an adaptor protein, our data open new insights into signaling events that are disrupted during transformation by PML-RARalpha and induced by retinoic acid during de novo differentiation of acute promyelocytic leukemia cells.
Subject(s)
Leukemia, Promyelocytic, Acute/metabolism , Neoplasm Proteins/physiology , Oncogene Proteins, Fusion/physiology , Proteins/metabolism , Tretinoin/pharmacology , Adaptor Proteins, Signal Transducing , Amino Acid Sequence , Base Sequence , Cell Differentiation , Cloning, Molecular , DNA, Complementary , Gene Expression Regulation, Neoplastic/drug effects , Humans , Leukemia, Promyelocytic, Acute/pathology , Molecular Sequence Data , Proteins/chemistry , Proteins/genetics , RNA, Messenger/genetics , Tumor Cells, Cultured , U937 CellsABSTRACT
CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leucoencephalopathy) is a small-artery disease of the brain caused by NOTCH3 mutations that lead to an abnormal accumulation of NOTCH3 within the vasculature. We aimed to establish whether immunostaining skin biopsy samples with a monoclonal antibody specific for NOTCH3 could form the basis of a reliable and easy diagnostic test. We compared the sensitivity and specificity of this method in two groups of patients suspected of having CADASIL with complete scanning of mutation-causing exons of NOTCH3 (in a retrospective series of 39 patients) and with limited scanning of four exons that are mutation hotspots (prospective series of 42 patients). In the retrospective series skin biopsy was positive in 21 (96%) of the 22 CADASIL patients examined and negative in all others; in the prospective series, seven of the 42 patients had a positive skin biopsy whereas only four had a mutation detected by limited NOTCH3 scanning. Our immunostaining technique is highly sensitive (96%) and specific (100%) for diagnosis of CADASIL.