Protein Stability and Transcription Factor Complex Assembly Determined by the SCL-LMO2 Interaction

Abstract

Gene expression programs are established by networks of interacting transcription factors. The basic helix-loop-helix factor SCL and the LIM-only protein LMO2 are components of transcription factor complexes that are essential for hematopoiesis. Here we show that LMO2 and SCL are predominant interaction partners in hematopoietic cells and that this interaction occurs through a conserved interface residing in the loop and helix 2 of SCL. This interaction nucleates the assembly of SCL complexes on DNA and is required for target gene induction and for the stimulation of erythroid and megakaryocytic differentiation. We also demonstrate that SCL determines LMO2 protein levels in hematopoietic cells and reveal that interaction with SCL prevents LMO2 degradation by the proteasome. We propose that the SCL-LMO2 interaction couples protein stabilization with higher order protein complex assembly, thus providing a powerful means of modulating the stoichiometry and spatiotemporal activity of SCL complexes. This interaction likely provides a rate-limiting step in the transcriptional control of hematopoiesis and leukemia, and similar mechanisms may operate to control the assembly of diverse protein modules.

High-throughput screening in niche-based assay identifies compounds to target preleukemic stem cells

“…Since pre-LSC self-renewal activity depends on the capacity of SCL to activate transcription (14), we next addressed the question of whether 2-ME2 inhibits SCL transcription activity, using a quantitative reporter assay (39,40). We observed that 2-ME2 but not DEXA inhibited SCL activity in transient transcription assays (Figure 4B), consistent with the suppression of endogenous SCL target genes in pre-LSCs ( Figure 4A).…”

The LMO2 oncogene regulates DNA replication in hematopoietic cells

“…LMO2 has a well-established function in transcriptional regulation via direct interaction with transcription factors, mostly of the bHLH family, SCL/TAL1, TAL2, and LYL1 or GATA proteins (2)(3)(4). In this study, we revealed unexpected new functions of LMO2 in hematopoiesis and leukemogenesis through a yeast two-hybrid screen of LMO2 interaction partners in hematopoietic progenitors.…”

“…We next addressed the question of whether LMO2 associates with the pre-RC in hematopoietic cells. Because pre-RC formation often requires a DNA template (35), we prepared DNA-containing chromatin extracts from CD34 + Kit + progenitors (TF-1 cells) (3,13,36,37) to assess the interaction of LMO2 with endogenous replication proteins by immunoprecipitation. We found that POLD1, PRIM1, and MCM6, but not lamin B or beta-actin (negative controls), reproducibly and specifically coimmunoprecipitate with LMO2, but not with control Ig, in TF-1 chromatin extracts (Fig.…”

Genetic interaction between Kit and Scl

“…50 We have expressed human SCL in mouse bone marrow cells because human and mouse SCL proteins are identical in their bHLH domains and more than 90% identical in their Nt domains. 51 Strikingly, the basic domain is conserved in Drosophila with only 2 conservative changes, I190V and S195T. This phylogenetic conservation underscores the importance of the basic domain for SCL function.…”