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. 2016 Aug 20;5:e17877. doi: 10.7554/eLife.17877

Figure 5. Exosome complex requirement for Kit expression.

Figure 5.

(A) Left: surface Kit in R1-R3 populations 48 hr post-Exosc8 knockdown. Representative plots. Right: surface Kit MFI relative to control R1 (mean ± SE, 5 biological replicates). (B) Left: surface Kit in R1-R3 cells 48 hr post-Exosc9 knockdown. Representative plots. Right: surface Kit MFI relative to control R1 (mean ± SE, 8 biological replicates). (C) Left: real-time RT-PCR of Kit mRNA and primary transcripts in sorted R1-R3 populations 72 hr post-infection with shControl or shExosc8 normalized to 18S and relative to control R1 (mean ± SE, 6 biological replicates). Middle: Kit Western blot with Ter119- cells 24 hr post-infection (mean ± SE, 3 biological replicates). (D) Left: real-time RT-PCR of erythroid mRNAs in sorted R1-R3 populations 72 hr post-infection with shControl or shExosc8 (mean ± SE, 6 biological replicates). Middle: GATA-2 and GATA-1 Western blot with Ter119- cells 24 hr post-infection. Right: densitometric analysis normalized to tubulin and relative to shControl (mean ± SE, 3 biological replicates). (E) qRT-PCR of Exosc8 and Kit mRNA and GATA-1/Exosc8-regulated cell cycle arrest genes in primary erythroid precursor cells 24 hr post-infection. Normalized to 18S and relative to the control (mean ± SE, 5 biological replicates). (F) Cell cycle analysis of control and Exosc8-knockdown Ter119- cells 24 (top) and 72 hr (bottom) post-infection (mean ± SE, 6 biological replicates) (G) qRT-PCR analysis of Exosc8 and Kit mRNA in G1E cells 48 hr post-infection with shControl or shExosc8 retrovirus, normalized to 18S and expressed relative to the control (mean ± SE, 3 independent experiments) (H) Cell surface Kit expression in infected (GFP+) and uninfected (GFP-) populations of G1E cells 48 hr post-infection with shExosc8 (mean ± SE, 3 independent experiments). Statistical analysis of control and treatment conditions was conducted with the Student’s T-test *p<0.05, **p<0.01, ***p<0.001. Source data is available in Figure 5—source data 1.

DOI: http://dx.doi.org/10.7554/eLife.17877.014

Figure 5—source data 1. This Excel spreadsheet contains the values for each biological replicate for data presented as either line graphs or histograms (mean ± SE) in Figure 5.
Sheet 1: Figure 5A Kit MFI in the R1, R2, R3, R4 and R5 population 48 hr after Exosc8 knockdown. Sheet 2: Figure 5B Kit MFI in the R1, R2, R3, R4 and R5 population 48 hr post-Exosc9 knockdown. Sheet 3: Figure 5C Kit mRNA and primary transcript expression sorted R1-R3 populations 72 hr post-infection with shControl or shExosc8 normalized to 18S and densitometry analysis of Kit protein in Ter119- cells 24 hr post-knockdown. Sheet 4: Figure 5D mRNA expression of erythroid genes in sorted R1-R3 populations 72 hr post-infection with shControl or shExosc8 and densitometry analysis of GATA-1 and GATA-2 protein Ter119- cells 24 hr post-knockdown. Sheet 5: Figure 5E Expression of Exosc8, Kit and GATA-1/Exosc8-regulated cell cycle arrest genes in primary erythroid precursor cells 24 hr post-infection, normalized to 18S. Sheet 6: Figure 5F Cell cycle analysis of control and Exosc8-knockdown Ter119− cells 24 and 72 hr post-infection. Sheet 7: Figure 5G Exosc8 and Kit mRNA expression in G1E cells 48 hr post-infection with shControl or shExosc8 retrovirus, normalized to 18S. Sheet 8: Figure 5H Kit MFI in infected (GFP+) and uninfected (GFP-) populations of G1E cells 48 hr post-infection with shExosc8.
DOI: http://dx.doi.org/10.7554/eLife.17877.015
elife-17877-fig5-data1.xlsx (65.9KB, xlsx)
DOI: 10.7554/eLife.17877.015