R40 - CSRP1 Knockdown Drives EMT-Associated Actin Remodeling in H1299 Lung Cancer Cells

Friday, October 10, 2025

10:00 AM - 11:00 AM PDT

Location: Exhibit Hall F, G & H

Presenting Author(s)

AB

Amrita K. Bedi

Undergraduate Student
University of California, Irvine
Mountain House, California, United States

Co-Author(s)

AB

Amrita K. Bedi

Undergraduate Student
University of California, Irvine
Mountain House, California, United States
Daisy Cortez

Undergraduate
University of California, Irvine
Santa Ana, California, United States
Ashley Urrutia Avila

PhD Candidate
University of California Irvine, United States
Timothy L. Downing

Associate Professor
University of California Irvine
Irvine, California, United States

Last Author(s)

Timothy L. Downing

Associate Professor
University of California Irvine
Irvine, California, United States

Co-Author(s)

MD

Michelle A. Digman

Professor
University of California Irvine, United States

Primary Investigator(s)

MD

Michelle A. Digman

Professor
University of California Irvine, United States

Introduction: : Lung cancer remains the leading cause of cancer-related mortality worldwide, primarily due to its high metastatic potential. The adhesome—a network of structural and signaling proteins that regulate cell adhesion, polarity, and cytoskeletal dynamics—plays a central role in tumor progression and metastasis. Dysregulation of adhesome components has been associated with poorer patient outcomes and reduced survival rates, highlighting its clinical relevance. In accordance with this, our lab recently demonstrated that knockdown of CSRP1 (CSRP1KD), an adhesome protein, enhances invasive and metastatic behavior in the H1299 lung cancer cell line.

To elucidate the cellular mechanisms driving this phenotype, we examined actin cytoskeletal organization and cell morphology, which are key features of epithelial-to-mesenchymal transition (EMT)—a critical step in cancer metastasis. EMT involves a loss of epithelial polarity and adhesion, accompanied by increased motility and invasiveness. Morphologically, EMT is associated with larger, more elongated cells, and reorganization of cortical actin into aligned stress fibers that facilitate migration. Based on these insights, we hypothesized that CSRP1KD in H1299 cells would trigger EMT-like actin restructuring, promoting a mesenchymal phenotype conducive to metastasis.

Materials and

Methods: : To assess actin reorganization indicative of EMT, H1299 scramble control and CSRP1KD cells were stained with fluorescent phalloidin, which specifically binds filamentous actin (F-actin). Image analysis quantified three key metrics: the Orientation Order Parameter (OOP) for actin alignment, cell area, and cell aspect ratio to evaluate cell size and elongation.

Results, Conclusions, and Discussions:: Scramble control cells displayed a low OOP, reflecting a diffused, disorganized actin network, whereas CSRP1KD cells exhibited aligned, stress fiber–like structures and markedly higher OOP. Additionally, CSRP1KD cells showed a significant increase in cell area and aspect ratio compared to controls, indicating a more elongated, mesenchymal morphology.

Collectively, these data suggest that CSRP1KD induces actin cytoskeletal remodeling consistent with EMT. Since actin reorganization drives cell migration and metastasis, this cytoskeletal shift likely underlies the increased invasive potential we observed.

Our findings indicate that CSRP1 knockdown in H1299 lung cancer cells induces actin cytoskeletal remodeling and morphological changes indicative of EMT. Specifically, CSRP1KD cells exhibited significantly greater actin fiber alignment (higher OOP), increased cell area, and elongation—hallmarks of transition toward a mesenchymal phenotype, which is associated with enhanced motility and metastatic potential. Future directions include validating EMT at the molecular and functional level. We will assess classic EMT markers including E‑cadherin, N‑cadherin, vimentin, and transcription factors such as Snail and ZEB1 to determine if the morphological shift reflects transcriptional EMT activation. These studies will clarify CSRP1’s contribution to actin-driven EMT and metastasis in lung cancer.

Acknowledgements and/or References (Optional): :