Lysyl hydroxylase 2 induces a collagen cross-link switch in tumor stroma
Yulong Chen, … , Mitsuo Yamauchi, Jonathan M. Kurie
Yulong Chen, … , Mitsuo Yamauchi, Jonathan M. Kurie
Published February 9, 2015
Citation Information: J Clin Invest. 2015;125(3):1147-1162. https://doi.org/10.1172/JCI74725.
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Research Article Oncology

Lysyl hydroxylase 2 induces a collagen cross-link switch in tumor stroma

Abstract

Epithelial tumor metastasis is preceded by an accumulation of collagen cross-links that heighten stromal stiffness and stimulate the invasive properties of tumor cells. However, the biochemical nature of collagen cross-links in cancer is still unclear. Here, we postulated that epithelial tumorigenesis is accompanied by changes in the biochemical type of collagen cross-links. Utilizing resected human lung cancer tissues and a p21CIP1/WAF1-deficient, K-rasG12D-expressing murine metastatic lung cancer model, we showed that, relative to normal lung tissues, tumor stroma contains higher levels of hydroxylysine aldehyde–derived collagen cross-links (HLCCs) and lower levels of lysine aldehyde–derived cross-links (LCCs), which are the predominant types of collagen cross-links in skeletal tissues and soft tissues, respectively. Gain- and loss-of-function studies in tumor cells showed that lysyl hydroxylase 2 (LH2), which hydroxylates telopeptidyl lysine residues on collagen, shifted the tumor stroma toward a high-HLCC, low-LCC state, increased tumor stiffness, and enhanced tumor cell invasion and metastasis. Together, our data indicate that LH2 enhances the metastatic properties of tumor cells and functions as a regulatory switch that controls the relative abundance of biochemically distinct types of collagen cross-links in the tumor stroma.

Authors

Yulong Chen, Masahiko Terajima, Yanan Yang, Li Sun, Young-Ho Ahn, Daniela Pankova, Daniel S. Puperi, Takeshi Watanabe, Min P. Kim, Shanda H. Blackmon, Jaime Rodriguez, Hui Liu, Carmen Behrens, Ignacio I. Wistuba, Rosalba Minelli, Kenneth L. Scott, Johannah Sanchez-Adams, Farshid Guilak, Debananda Pati, Nishan Thilaganathan, Alan R. Burns, Chad J. Creighton, Elisabeth D. Martinez, Tomasz Zal, K. Jane Grande-Allen, Mitsuo Yamauchi, Jonathan M. Kurie

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Figure 3

LH2 expression is upregulated in human and murine lung cancer and promotes the migration and invasion of KC cells.

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LH2 expression is upregulated in human and murine lung cancer and promot...
(A) The 67 genes differentially expressed between highly and poorly metastatic KC cells (KC1–KC3 versus KC4) with the greatest fold change (up- or downregulation) were analyzed in 15 human lung cancer cohorts (https://www.oncomine.org). For each gene (dot), location on the y axis indicates fold difference between the two groups of murine cells; location on the x axis indicates the number of human data sets in which the gene is differentially expressed in cancer relative to normal lung (P < 0.01, >1.5-fold). (B) LH2 mRNA (bar graph) and protein (gels) levels measured using Q-PCR (triplicate samples) and immunoblotting, respectively. P < 0.0001 (1-way ANOVA). (C) LH2a and LH2b isoforms measured by RT-PCR. (D) Q-PCR analysis (bar graphs) and immunoblot analysis (gels) of KC2 cells stably transfected with vectors expressing LH2 shRNA (SH1, SH3, and SH5) or scrambled control shRNA (SCR). Normalized Q-PCR results are expressed as mean ± SD values of triplicate samples. (E) Images show migrating and invading cells in Boyden chambers. Scale bars: 200 μm. Bar graphs show mean ± SD of triplicate wells expressed as the percent change relative to control transfectants (SCR), which were set at 100%. (F) Scatter plots of primary tumor weights (left) and total numbers of lung metastases (right). Dot, single mouse; bars and whiskers, mean ± SD. P values shown in figure, 2-tailed Student’s t test. L32 and actin were used as loading controls for Q-PCR and Western blot assays, respectively. Each result is from a single experiment.