TAL1 and LIM-Only Proteins Synergistically Induce Retinaldehyde Dehydrogenase 2 Expression in T-Cell Acute Lymphoblastic Leukemia by Acting as Cofactors for GATA3

Abstract

Previously, we have shown that TAL1 and the LIM-only protein gene (LMO) are regularly coactivated in T-cell acute lymphoblastic leukemia (T-ALL). This observation is likely to relate to the findings that TAL1 and LMO are highly synergistic in T-cell tumorigenesis in double-transgenic mice. To understand the molecular mechanisms of functional synergy between TAL1 and LMO in tumorigenesis and transcriptional regulation, we tried to identify downstream target genes regulated by TAL1 and LMO by a subtractive PCR method. One of the isolated genes, that for retinaldehyde dehydrogenase 2 (RALDH2), was regularly expressed in most of the T-ALL cell lines that coexpressed TAL1 and LMO. Exogenously transfected TAL1 and LMO, but not either alone, induced RALDH2 expression in a T-ALL cell line, HPB-ALL, not expressing endogeneous TAL1 or LMO. The RALDH2 transcripts in T-ALL were, however, mostly initiated within the second intron. Promoter analysis revealed that a GATA site in a cryptic promoter in the second intron was essential and sufficient for the TAL1-and LMO-dependent transcriptional activation, and GATA3 binds to this site. In addition, forced expression of GATA3 potentiated the induction of RALDH2 by TAL1 and LMO, and these three factors formed a complex in vivo. Furthermore, a TAL1 mutant not binding to DNA also activated the transcription of RALDH2 in the presence of LMO and GATA3. Collectively, we have identified the RALDH2 gene as a first example of direct transcriptional target genes regulated by TAL1 and LMO in T-ALL. In this case, TAL1 and LMO act as cofactors for GATA3 to activate the transcription of RALDH2.In various types of leukemia, specific recurrent chromosomal translocations are frequently observed and potential oncogenic transcription factors have been identified from the chromosomal breakpoints (46). These transcription factors are mostly involved in normal hematopoietic cell differentiation and growth. In T-cell acute lymphoblastic leukemia (T-ALL), frequent chromosomal abnormalities are found in the TAL1 (also called SCL or TCL5) locus (5, 9, 10, 14). Ectopic expression of TAL1, which is not normally expressed in T cells (44,57), is observed in ϳ60% of T-ALL patients (4). TAL1 encodes at least two alternative isoforms, full-length TAL1␣ and Nterminally truncated TAL1␤, having a basic helix-loop-helix (bHLH) motif found in a number of transcription factors involved in the regulation of cell differentiation (6). TAL1 dimerizes with ubiquitous bHLH E-proteins (E47, E12, and HEB) (20,22), and the heterodimers bind to the E-box motif (CANNTG), most preferably to AACAGATGGT (21). However, no downstream target genes have been identified. It is thus uncertain whether TAL1-E-protein heterodimers regulate transcription through binding to this preferred E-box.LMO1 (RBTN1 or TTG1) and LMO2 (RBTN2 or TTG2) are also genes originally identified from recurrent chromosomal breakpoints in T-ALL (7,34,49). Their expression in T-ALL is ectopic, like that of TAL1 (15,18,35,49). They encode highly related LIM-only cla...

Expression of an engrailed-LMO4 fusion protein in mammary epithelial cells inhibits mammary gland development in mice

“…LMO proteins do not bind DNA directly, but regulate gene transcription by associating with other transcription factors. This model is supported by studies showing that LMOs, through their LIM domains, exist in a stable complex with helix loop helix (HLH) heterodimeric partner proteins that include TAL1(SCL)/E12 (Valge-Archer et al, 1994;Wadman et al, 1994Wadman et al, , 1997Osada et al, 1995Osada et al, , 1997Larson et al, 1996;Visvader et al, 1997;Ono et al, 1998;Bao et al, 2000;Herblot et al, 2000;Mead et al, 2001), and GATA factors (Osada et al, 1995;Wadman et al, 1997;Ono et al, 1998;Mead et al, 2001). In addition, LIM domains of the LIM homeodomain and LMO proteins interact strongly with cofactors, including the coactivators Clim1 and Clim2/Ldb1/NLI (Agulnick et al, 1996;Jurata et al, 1996;Bach et al, 1997Bach et al, , 1999Visvader et al, 1997), which confer transcriptional activation and promote synergism between DNA-binding proteins (Bach, 2000).…”

GATA3 controls the expression of CD5 and the T cell receptor during CD4 T cell lineage development

“…Although by RT-PCR and DNA micro-array analyses we did not detect significant differences in E2A transcript levels between CD2-GATA3 and WT DP cells (M. de Bruijn, unpublished), it is very well possible that GATA3 acts by stabilizing E2A protein levels, as in erythrocytes E2A has been shown to be part of a large DNA-binding complex containing the GATA family member GATA1 [33], and the related helix-loophelix transcription factor Tal-1/Scl has been shown to interact directly with GATA1 and GATA3 [34,35]. A role for GATA3 as a negative regulator of CD5 gene expression would also be supported by our observation in micro-array experiments that enforced GATA3 expression is associated with down-regulation of CD6 (J. P. van Hamburg et al, unpublished), which is closely linked to CD5 in the genome and has a similar pattern of expression ( [31] and references therein).…”

Helt, a Novel Basic-Helix-Loop-Helix Transcriptional Repressor Expressed in the Developing Central Nervous System

“…Total RNA was isolated using an RNeasy mini kit (Qiagen). Subtractive PCR was performed as described (33). After four rounds of subtractive hybridization, the amplified cDNA fragments were cloned and sequenced.…”

“…293E cells (2 ϫ 10 6 ) were transfected with the indicated combinations of 5 g of each plasmid using TransIT LT1 reagent (Mirus Corp.). Immunoprecipitation experiments and detection by Western blotting were performed as described (33).…”