Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2021 Apr 9;6:144. doi: 10.1038/s41392-021-00534-2

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

Fig. 6 — KRAS drives the production of CSF2 and lactate in tumor cells by stabilizing HIF-1α. a Representative immunoblot for HIF-1α in whole-cell lysates from KRAS mutant and wild-type CRC cells cultured at 21% O₂. b Representative immunoblot for HIF-1α in SW48 cells expressing SW48-Vector or SW48-KRAS^G12V cultured at 1% O₂ for the indicated times. c Representative immunoblot of the expression kinetics of HIF-1α in MOCK (SW620-siMOCK) or KRAS siRNA-transfected (SW620-siKRAS) SW620 cells treated with the transcription inhibitor cycloheximide (CHX, 100 μg/ml) for the indicated times at 21% O₂. d HIF-1α target genes were measured by qRT-PCR (n = 3) in the indicated cells cultured under normoxia. e Representative immunoblot of the expression of HIF-1α, hydroxy-HIF-1α, PHD2, and VHL in MOCK or KRAS siRNA-transfected SW620 cells cultured at 21% O₂. Before protein extraction, cells were treated with a proteasome inhibitor (10 μM MG-132) for 1 h to demonstrate hydroxyl-HIF-1α. f Immunoblots for HIF-1α in SW48-Vector or SW48-KRAS^G12V cells treated with 1 mM DMOG for the indicated times at 21% O₂. g Fold change (FC) of HIF-1α target genes (GLUT1, HK2, CSF2) in response to hypoxia (n = 3) measured by qRT-PCR. The ratio of hypoxic to normoxic gene expression is shown. h FC of HIF-1α target genes in response to DMOG treatment was measured by qRT-PCR and the ratio of untreated to DMOG-treated gene expression is shown (n = 3). i Immunoblots of SW48-Vector or SW48-KRAS^G12V cells incubated with 10 mM N-acetylcysteine (NAC) and cultured at 1% O₂. j Immunoblots of SW48-Vector or SW48-KRAS^G12V cells cultured at 21% O₂ with 10 mM NAC or 1 mM DMOG as indicated. k Schematic summarizing our proposed model for the crosstalk between KRAS and TAMs in CRC. KRAS drives the production of CSF2 and lactate in tumor cells by stabilizing HIF-1α, and CSF2 synergizes with lactate to induce functional reprogramming of TAMs, which in turn supports tumor progression. β-actin served as loading controls and 3 independent experiments were performed and yielded similar results (a–c, e, f, i, j). β-actin serves as a housekeeping gene for qRT-PCR (d, g, h). *p ≤ 0.05 and **p ≤ 0.01, by two-tailed Student’s t-test (d, g, h)