MEIS-mediated suppression of human prostate cancer growth and metastasis through HOXB13-dependent regulation of proteoglycans
- The Committee on Cancer Biology, The University of Chicago, United States
- Department of Pathology, The University of Illinois at Chicago, United States
- Department of Surgery, Section of Urology, The University of Chicago, United States
- Department of Urology, UT Southwestern, United States
Figures
Figure 1 with 2 supplements
Expression of MEIS1 or MEIS2 in PrCa cell lines is sufficient to decrease growth in vitro and in vivo.
(A) RT-PCR for MEIS1 (black) or pan-MEIS2 (gray) in five of the most common prostate cancer cell lines (LNCaP, CWR22Rv1, LAPC4, Du145, and PC3) as compared to non-malignant primary prostate …
Figure 1—figure supplement 1
MEIS2 Isoforms in Prostate Cancer Cells.
(A) Schematic indicating the most common translational isoforms of MEIS2 (UniProt Identifier #O14770) modified from Geerts et al., 2005. Boxes represent exons as numbered on top. The checkered box …
Figure 1—figure supplement 2
Decreased cell number with exogenous MEIS1 or MEIS2 expression is not the result of increased cell death.
Representative images (left) and quantitation (right) of terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labeling (TUNEL) assay on CWR22Rv1 and LAPC4 cells expressing control, LV-MEIS1, or …
Figure 2
Exogenous MEIS1 expression rescues the nuclear MEIS1–HOXB13 interaction present in normal prostate epithelial cells.
(A) Heatmap for HOX expression profile from publicly available, human RNA-seq data of benign prostate, primary prostate tumor, or metastasis from a prostate tumor. Log2(FPKM) values were used to …
Figure 3 with 1 supplement
MEIS-mediated suppression of proliferation and migration is dependent upon HOXB13.
(A) Western blot analysis of HOXB13 knockout using CRISPR and exogenous expression of MEIS1 in the resulting HOXB13ko cells for CWR22Rv1 and LAPC4. Actin was used as a loading control. (B-C) …
Figure 3—figure supplement 1
Cell viability is unaffected by HOXB13ko in LAPC4 cells.
Cell viability over time of LAPC4-Control, -LV-MEIS1, -HOXB13ko, and -HOXB13ko-LV-MEIS1.
Figure 4
MEIS-mediated metastasis suppression in vivo is HOXB13-dependent.
(A) Schematic of experimental design for intracardiac injection of CWR22Rv1 cell line derivatives into athymic nude mice to model metastasis. Control (black), LV-MEIS1 (blue), HOXB13ko (green), and …
Figure 5 with 2 supplements
Integration of ChIP-seq and RNA-seq analyses reveals MEIS1-mediated, HOXB13-dependent, direct regulation of proteoglycans including Decorin (DCN).
(A) Western blot confirmation of pull-down of MEIS1 from chromatin in CWR22Rv1 LV-MEIS1 and HOXB13ko-LV-MEIS1 cell lines (Top) and heatmap of read density profiles from MEIS1 ChIP-seq for ±2 kb of …
Figure 5—figure supplement 1
HOXA9 motif shares similarity, RNA-seq MDS plot, and Validation of DCN expression in VCaP.
(A) The web-based tool STAMP for alignment, similarity, and database-matching of DNA motifs was used to identify other motifs that share similarity to the HOXA9 motif inferred in SpaMo analysis of …
Figure 5—figure supplement 2
Functional impact of MEIS1 and DCN knockdown in LAPC4.
Western blot of DCN demonstrating decreased DCN protein expression in LAPC4 cells when MEIS1, MEIS2, or both MEIS1 and MEIS2 are depleted using targeted shRNAs. These lines were originally reported …
Figure 6 with 1 supplement
Knockdown of DCN partially reverses MEIS-mediated tumor suppression.
(A) RT-PCR in both CWR22Rv1 and LAPC4 for DCN expression in control (black) and LV-MEIS1 (dark blue) cells treated with a non-silencing control siRNA pool (siNSC, 4 siRNAs in pool) or LV-MEIS1 cells …
Figure 6—figure supplement 1
Western blot analysis of MEIS1, HOXB13, DCN, and known downstream targets of DCN in LAPC4 control-siNSC, LV-MEIS1-siNSC, and LV-MEIS1-siDCN cells.
Figure 7
Increased proteoglycan expression within human prostate tumors that retain MEIS1/2 expression.
(A) Western blot analysis of MEIS1 and DCN expression in protein lysates from primary prostate epithelial cells (PrECs). Actin was used as a loading control. (B) Correlation of mRNA expression level …
Author response image 1
This file contains the HOXB13 binding to DCN and LUM of every publicly available HOXB13 ChIP-seq experiment done in prostate tissue or cell lines.
Every column is a different study or different treatment condition from the same study. The SRX###### number at the top of each column to be taken to more info about the study/experimental design …
Additional files
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Supplementary file 1
MEIS1 ChIP-seq peaks and annotation in CWR22Rv1 LV-MEIS1, Related to Figure 5.
Peaks identified by MACS2 in LV-MEIS1 ChIP-seq samples vs. input DNA were annotated to the nearest transcription start site (TSS) using HOMER.
- https://cdn.elifesciences.org/articles/53600/elife-53600-supp1-v2.xlsx
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Supplementary file 2
MEIS1 ChIP-seq peaks and annotation in CWR22Rv1 HOXB13ko-LV-MEIS1, Related to Figure 5.
Peaks identified by MACS2 in HOXB13ko-LV-MEIS1ChIP-seq samples vs. input DNA were annotated to the nearest transcription start site (TSS) using HOMER.
- https://cdn.elifesciences.org/articles/53600/elife-53600-supp2-v2.xlsx
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Supplementary file 3
Counts per million (CPM) expression of genes in RNA-seq from CWR22Rv1 cell line derivatives, Related to Figure 5.
Adapters and low-quality reads were trimmed before aligning sequences to hg38 transcriptome (ENSEMBL) using kallisto. Transcript counts were then summarized to the gene level with tximport and further analyzed with EdgeR to remove genes with low counts and normalize to library sizes.
- https://cdn.elifesciences.org/articles/53600/elife-53600-supp3-v2.xlsx
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Supplementary file 4
All significantly differentially expressed genes between CWR22Rv1-LV-MEIS1 and control cells, Related to Figure 5.
TREAT and GLM methodologies in edgeR were used to determine significantly differentially expressed genes (fold-change >±1.5, FDR < 0.05) in LV-MEIS1 vs. control cells.
- https://cdn.elifesciences.org/articles/53600/elife-53600-supp4-v2.xlsx
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Supplementary file 5
DEGs that are direct targets of MEIS1 only when HOXB13 is present, Related to Figure 5.
Overlap of DEGs between LV-MEIS1 and control cells with ChIP-seq targets from both HOXB13ko and HOXB13ko-LV-MEIS1 cells identified 157 DEGs that are targets of MEIS1 only when HOXB13 is present.
- https://cdn.elifesciences.org/articles/53600/elife-53600-supp5-v2.xlsx
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Supplementary file 6
GSEA for Gene Ontology: Biological Processes on RNA-seq from LV-MEIS1 and control cells, Related to Figure 5.
The top 20 enriched gene sets from CWR22Rv1 RNA-seq of GSEA on the Gene Ontology: Biological Processes collection from MSigDB.
- https://cdn.elifesciences.org/articles/53600/elife-53600-supp6-v2.xlsx
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Supplementary file 7
Key resources table.
- https://cdn.elifesciences.org/articles/53600/elife-53600-supp7-v2.docx
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Transparent reporting form
- https://cdn.elifesciences.org/articles/53600/elife-53600-transrepform-v2.pdf
