Abstract
Elucidation of the molecular mechanisms that are responsible for regulating the most basic properties of stem cells, self-renewal, and differentiation remains a major challenge in hematopoietic stem cell biology. We have taken a functional genomics approach towards revealing these mechanisms. Previous studies of the fetal liver genetic program led to the development of Stem Cell Database (SCDb, http://stemcell.princeton.edu), a resource for the stem cell community. These studies have been expanded to include the microenvironmental component of hematopoiesis and are the focus herein. In our efforts to study the microenvironmental component we have identified a stromal cell line, AFT024, which serves as a surrogate stem cell niche. The line provides a milieu that facilitates the maintenance of transplantable mouse and human stem cells as well as the generation of large populations of committed progenitors. In a manner mirroring the work done with the SCDb, we provide an online resource, Stromal Cell Database, StroCDB (http://stromalcell.princeton.edu), that is a compendium of information and data derived from biological and molecular studies of this surrogate niche. These include bioinformatic analyses of over 6000 clones derived from a subtracted library enriched for messages expressed in AFT024 as well as data derived from custom expression arrays developed from this library. Herein we describe these efforts and provide a guide for navigating the database and mining the information contained within.
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References
Phillips, R. L., Ernst, R. E., Brunk, B., Ivanova, N., Mahan, M. A., Deanehan, J. K., et al. (2000) The genetic program of hematopoietic stem cells. Science 288, 1635–1640.
Ivanova, N. B., Dimos, J. T., Schaniel, C., Hackney, J. A., Moore, K. A., and Lemischka, I. R. (2002) A stem cell molecular signature. Science 298, 601–604.
Lemischka, I. (2001) Stem cell dogmas in the genomics era. Rev. Clin. Exp. Hematol. 5, 15–25.
Lemischka, I. (1999) Searching for stem cell regulatory molecules. Some general thoughts and possible approaches. Ann. N Y Acad. Sci. 872, 274–287; discussion 287–288.
Wineman, J., Moore, K., Lemischka, I., and Muller-Sieburg, C. (1996) Functional heterogeneity of the hematopoietic microenvironment: rare stromal elements maintain long-term repopulating stem cells. Blood 87, 4082–4090.
Moore, K. A., Ema, H., and Lemischka, I. R. (1997) In vitro maintenance of highly purified, transplantable hematopoietic stem cells. Blood 89, 4337–4347.
Thiemann, F. T., Moore, K. A., Smogorzewska, E. M., Lemischka, I. R., and Crooks, G. M. (1998) The murine stromal cell line AFT024 acts specifically on human CD34+CD38-progenitors to maintain primitive function and immunophenotype in vitro. Exp. Hematol. 26, 612–619.
Miller, J. S., McCullar, V., Punzel, M., Lemischka, I. R., and Moore, K. A. (1999) Single adult human CD34(+)/Lin-/CD38(−) progenitors give rise to natural killer cells, B-lineage cells, dendritic cells, and myeloid cells. Blood 93, 96–106.
Punzel, M., Wissink, S. D., Miller, J. S., Moore, K. A., Lemischka, I. R., and Verfaillie, C. M. (1999) The myeloid-lymphoid initiating cell (ML-IC) assay assesses the fate of multipotent human progenitors in vitro. Blood 93, 3750–3756.
Lewis, I. D., Almeida-Porada, G., Du, J., Lemischka, I. R., Moore, K. A., Zanjani, E. D., et al. (2001) Umbilical cord blood cells capable of engrafting in primary, secondary, and tertiary xenogeneic hosts are preserved after ex vivo culture in a noncontact system. Blood 97, 3441–3449.
Nolta, J. A., Thiemann, F. T., Arakawa-Hoyt, J., Dao, M. A., Barsky, L. W., Moore, K. A., et al. (2002) The AFT024 stromal cell line supports long-term ex vivo maintenance of engrafting multipotent human hematopoietic progenitors. Leukemia 16, 352–361.
Hackney, J. A., Charbord, P., Brunk, B. P., Stoeckert, C. J., Lemischka, I. R., and Moore, K. A. (2002) A molecular profile of a hematopoietic stem cell niche. Proc. Natl. Acad. Sci. USA 99, 13,061–13,066.
Li, W. B., Gruber, C. E., Lin, J. J., Lim, R., D’Alessio, J. M., and Jessee, J. A. (1994) The isolation of differentially expressed genes in fibroblast growth factor stimulated BC3H1 cells by subtractive hybridization. BioTechniques 16, 722–729.
Altschul, S. F., Madden, T. L., Schaffer, A. A., Zhang, J., Zhang, Z., Miller, W., et al. (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389–402.
Benson, D. A., Karsch-Mizrachi, I., Lipman, D. J., Ostell, J., Rapp, B. A., and Wheeler, D. L. (2002) GenBank. Nucleic Acids Res. 30, 17–20.
Boeckmann, B., Bairoch, A., Apweiler, R., Blatter, M. C., Estreicher, A., Gasteiger, E., et al. (2003) The SWISS-PROT protein knowledgebase and its supplement TrEMBL in 2003. Nucleic Acids Res. 31, 365–370.
The Computational Biology and Informatics Laboratory. DoTS: a database of transcribed sequences for human and mouse genes. Center for Bioinformatics, University of Pennsylvania. http://www.cbil.upenn.edu/downloads/DoTS/.
The Computational Biology and Informatics Laboratory. AllGenes: a web site providing access to an integrated database of known and predicted human and mouse genes. Version 5.0 Ed. Center for Bioinformatics, University of Pennsylvania. http://www.allgenes.org.
Ponting, C. P., Schultz, J., Milpetz, F., and Bork, P. (1999) SMART: identification and annotation of domains from signalling and extracellular protein sequences. Nucleic Acids Res. 27, 229–232.
Nielsen, H., Engelbrecht, J., Brunak, S., and von Heijne, G. (1997) Identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Protein Eng. 10, 1–6.
Huang, X. (1996) An improved sequence assembly program. Genomics 33, 21–31.
Moore, K. A., Pytowski, B., Witte, L., Hicklin, D., and Lemischka, I. R. (1997) Hematopoietic activity of a stromal cell transmembrane protein containing epidermal growth factor-like repeat motifs. Proc. Natl. Acad. Sci. USA 94, 4011–4016.
Tavazoie, S., Hughes, J. D., Campbell, M. J., Cho, R. J., and Church, G. M. (1999) Systematic determination of genetic network architecture. Nat. Genet. 22, 281–285.
Ploemacher, R. E., van der Sluijs, J. P., van Beurden, C. A., Baert, M. R., and Chan, P. L. (1991) Use of limiting-dilution type long-term marrow cultures in frequency analysis of marrow-repopulating and spleen colony-forming hematopoietic stem cells in the mouse. Blood 78, 2527–2533.
Churchill, G. A. (2002) Fundamentals of experimental design for cDNA microarrays. Nat. Genet. 32(Suppl), 490–495.
Quackenbush, J. (2002) Microarray data normalization and transformation. Nat. Genet. 32(Suppl), 496–501.
Slonim, D. K. (2002) From patterns to pathways: gene expression data analysis comes of age. Nat. Genet. 32(Suppl), 502–508.
Brazma, A., Hingamp, P., Quackenbush, J., Sherlock, G., Spellman, P., Stoeckert, C., et al. (2001) Minimum information about a microarray experiment (MIAME)-toward standards for microarray data. Nat. Genet. 29, 365–371.
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Hackney, J.A., Moore, K.A. (2005). A Functional Genomics Approach to Hematopoietic Stem Cell Regulation. In: Baron, M.H. (eds) Developmental Hematopoiesis. Methods in Molecular Medicine, vol 105. Humana Press, Totowa, NJ. https://doi.org/10.1385/1-59259-826-9:439
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DOI: https://doi.org/10.1385/1-59259-826-9:439
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