Abstract

The facilitating chromatin transcription (FACT) complex, a heterodimer of SSRP1 and Spt16, has been shown to regulate transcription elongation through a chromatin template in vitro and on specific genes in cells. However, its global role in transcription regulation in human cells remains largely elusive. We conducted spotted microarray analyses using arrays harboring 8308 human genes to assess the gene expression profile after knocking down SSRP1 or Spt16 levels in human non-small cell lung carcinoma (H1299) cells. Although the changes of these transcripts were surprisingly subtle, there were approximately 170 genes whose transcript levels were either reduced or induced >1.5-fold. Approximately 106 genes with >1.2-fold change at the level of transcripts were the common targets of both SSRP1 and Spt16 ( approximately 1.3%). A subset of genes was regulated by SSRP1 independent of Spt16. Further analyses of some of these genes not only verified this observation but also identified the serum-responsive gene, egr1, as a novel target for both SSRP1 and Spt16. We further showed that SSRP1 and Spt16 are important for the progression of elongation RNA pol II on the egr1 gene. These results suggest that SSRP1 has Spt16-dependent and -independent roles in regulating gene transcription in human cells.

Highlights

  • facilitating chromatin transcription (FACT) is a heterodimeric complex consisting of Spt16 and SSRP1 (8, 9)

  • It has been shown that the FACT complex functions in transcriptional elongation on a chromatin template in vitro and on specific genes in cells (7, 25)

  • As described here, indicates that SSRP1 and Spt16 share a number of common target genes (Figs. 2 and 3; Tables 2 and 3), their global roles in gene transcription are not apparent

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Summary

Introduction

FACT is a heterodimeric complex consisting of Spt and SSRP1 (structure-specific recognition protein-1) (8, 9). Human Spt is a 120-kDa protein and contains a highly acidic and serine-rich carboxyl terminus. It binds to H2AH2B dimers and to mononucleosomes (10). The yeast Spt16/Cdc was identified in two independent screens for genes involved in transcription regulation (11, 12). It remains obscure if SSRP1 has an Spt16-independent role in gene regulation It is still unclear if FACT plays a global or gene-specific role in transcriptional regulation in human cells. To address these questions, we generated tet-inducible siRNA cell lines for each of these two proteins using H1299 cells that are p53-deficient (31). Our study suggests that SSRP1 and Spt work together for the expression of a number of genes, whereas SSRP1 appears to have an independent role in regulating the expression of a subset of genes in human cells

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