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A small molecule Abl kinase inhibitor induces differentiation of Abelson virus–transformed pre-B cell lines

Abstract

Abelson murine leukemia virus–transformed cell lines have provided a critical model system for studying the regulation of B cell development. However, transformation by v-Abl blocks B cell development, resulting in the arrest of these transformants in an early pre-B cell–like state. We report here that treatment of Abelson virus–transformed pre-B cell lines with the small molecule Abl kinase inhibitor (STI571) results in their differentiation to a late pre-B cell–like state characterized by induction of immunoglobulin (Ig) light chain gene rearrangement. DNA microarray analyses enabled us to identify two genes inhibited by v-Abl that encode the Igk 3′ enhancer–binding transcription factors Spi-B and IRF-4. We show that enforced expression of these two factors is sufficient to induce germline Igk transcription in Abelson-transformed pro-B cell lines. This suggests a key role for these factors, and perhaps for c-Abl itself, in the regulated activation of Ig light chain gene rearrangement.

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Figure 1: STI571 induces germline Igk and Rag1 and Rag2 transcription in pro-B cell lines.
Figure 2: STI571 stimulates Ig light chain gene rearrangement.
Figure 3: STI571 treatment promotes the accessibility of light chain genes to RAG-1 and RAG-2–mediated RSS cleavage in vitro.
Figure 4: Results of an Affymetrix microarray screen for genes regulated by STI571.
Figure 5: Analysis of transcripts in 220-8 pro-B cells during a time-course of STI571 treatment.
Figure 6: Overexpression of Spib and Irf4 activates Igk germline transcription.
Figure 7: Quantitative analysis of RAG-1 and RAG-2, IRF-4 and Spi-B transcripts in various sorted primary pre-B cell fractions.

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Acknowledgements

We thank R.Tjian, R. Freiman, M. Holmes and Y. Isogai (HHMI, Berkeley) for use of and help with equipment required for the Affymetrix experiments; C. Lowell (UCSF) and C. Sawyers (UCLA) for assistance; N. Rosenberg, C. Guidos, F. Alt and D. Baltimore for cell lines; H. Singh (HHMI, Chicago) for Spi-B and IRF-4 cDNAs; L-Y. Hsu for designing the quantitative RT-PCR assay for Rag1 and Rag2 and J. Curry for help with the analysis; H. Nolla (Cancer Research Lab, UC Berkeley) for cell sorting; and members of the Schlissel Lab as well as A. Winoto and C. Thompson for useful comments on the manuscript. Supported by grants from the NIH (RO1 HL48702) and the Arthritis Foundation (to M.S.S.).

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Correspondence to Mark S. Schlissel.

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Supplementary information

Web Fig. 1.

Flow cytometric analyses of CD25/IL2Rα and CD62L/L-selectin cell surface expression in STI571-treated 220-8 pro-B cells. Cell surface expression of CD25 and CD62L were analyzed by flow cytometry using either PE-conjugated anti-CD25 or anti-CD62L monoclonal antibodies. The data were gated for live cells based on forward and side scatter. Top panel shows anti-CD25 staining of 220-8 cells treated with 10 μM STI571 for 12 hours (unshaded) overlaid with the histogram for untreated 220-8 (shaded dark). Bottom panel shows anti-CD62L staining of 220-8 cells treated with 10 μM STI571 for 12 hours (unshaded) overlaid with the histogram for untreated 220-8 (shaded dark). (PDF 19 kb)

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Muljo, S., Schlissel, M. A small molecule Abl kinase inhibitor induces differentiation of Abelson virus–transformed pre-B cell lines. Nat Immunol 4, 31–37 (2003). https://doi.org/10.1038/ni870

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