Analysis of gene expression profiles in HeLa cells in response to overexpression or siRNA-mediated depletion of NASP
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Analysis of gene expression profiles in HeLa cells in response to overexpression or siRNA-mediated depletion of NASP Oleg M Alekseev*, Richard T Richardson, Oleg Alekseev and Michael G O'Rand Address: Department of Cell and Developmental Biology, University of North Carolina at Chapel Hill, Chapel Hill, NC, 27599-7090, USA Email: Oleg M Alekseev* - [email protected]; Richard T Richardson - [email protected]; Oleg Alekseev - [email protected]; Michael G O'Rand - [email protected] * Corresponding author
Published: 13 May 2009 Reproductive Biology and Endocrinology 2009, 7:45
doi:10.1186/1477-7827-7-45
Received: 4 March 2009 Accepted: 13 May 2009
This article is available from: http://www.rbej.com/content/7/1/45 © 2009 Alekseev et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract Background: NASP (Nuclear Autoantigenic Sperm Protein) is a linker histone chaperone required for normal cell division. Changes in NASP expression significantly affect cell growth and development; loss of gene function results in embryonic lethality. However, the mechanism by which NASP exerts its effects in the cell cycle is not understood. To understand the pathways and networks that may involve NASP function, we evaluated gene expression in HeLa cells in which NASP was either overexpressed or depleted by siRNA. Methods: Total RNA from HeLa cells overexpressing NASP or depleted of NASP by siRNA treatment was converted to cRNA with incorporation of Cy5-CTP (experimental samples), or Cy3-CTP (control samples). The labeled cRNA samples were hybridized to whole human genome microarrays (Agilent Technologies, Wilmington, Delaware, USA). Various gene expression analysis techniques were employed: Significance Analysis of Microarrays (SAM), Expression Analysis Systematic Explorer (EASE), and Ingenuity Pathways Analysis (IPA). Results: From approximately 36 thousand genes present in a total human genome microarray, we identified a set of 47 up-regulated and 7 down-regulated genes as a result of NASP overexpression. Similarly we identified a set of 56 up-regulated and 71 down-regulated genes as a result of NASP siRNA treatment. Gene ontology, molecular network and canonical pathway analysis of NASP overexpression demonstrated that the most significant changes were in proteins participating in organismal injury, immune response, and cellular growth and cancer pathways (major "hubs": TNF, FOS, EGR1, NFκB, IRF7, STAT1, IL6). Depletion of NASP elicited the changed expression of proteins involved in DNA replication, repair and development, followed by reproductive system disease, and cancer and cell cycle pathways (major "hubs": E2F8, TP53, FGF, FSH, FST, hCG, NFκB, TRAF6). Conclusion: This study has demonstrated that NASP belongs to a network of g
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