Global Effects of DDX3 Inhibition on Cell Cycle Regulation Identified by a Combined Phosphoproteomics and Single Cell Tracking Approach

Marise R. Heerma van Voss, Kai Kammers, Farhad Vesuna, Justin Brilliant, Yehudit Bergman, Saritha Tantravedi, Xinyan Wu, Robert N Cole, Andrew Holland, Paul J. van Diest, Venu Raman

Research output: Contribution to journalArticle

Abstract

DDX3 is an RNA helicase with oncogenic properties. The small molecule inhibitor RK-33 is designed to fit into the ATP binding cleft of DDX3 and hereby block its activity. RK-33 has shown potent activity in preclinical cancer models. However, the mechanism behind the antineoplastic activity of RK-33 remains largely unknown. In this study we used a dual phosphoproteomic and single cell tracking approach to evaluate the effect of RK-33 on cancer cells. MDA-MB-435 cells were treated for 24 hours with RK-33 or vehicle control. Changes in phosphopeptide abundance were analyzed with quantitative mass spectrometry using isobaric mass tags (Tandem Mass Tags). At the proteome level we mainly observed changes in mitochondrial translation, cell division pathways and proteins related to cell cycle progression. Analysis of the phosphoproteome indicated decreased CDK1 activity after RK-33 treatment. To further evaluate the effect of DDX3 inhibition on cell cycle progression over time, we performed timelapse microscopy of Fluorescent Ubiquitin Cell Cycle Indicators labeled cells after RK-33 or siDDX3 exposure. Single cell tracking indicated that DDX3 inhibition resulted in a global delay in cell cycle progression in interphase and mitosis. In addition, we observed an increase in endoreduplication. Overall, we conclude that DDX3 inhibition affects cells in all phases and causes a global cell cycle progression delay.

Original languageEnglish (US)
Pages (from-to)755-763
Number of pages9
JournalTranslational Oncology
Volume11
Issue number3
DOIs
StatePublished - Jun 1 2018

Fingerprint

Cell Tracking
Cell Cycle
Endoreduplication
RNA Helicases
Phosphopeptides
Interphase
Proteome
Ubiquitin
Mitosis
Cell Division
Antineoplastic Agents
Microscopy
Mass Spectrometry
Neoplasms
Adenosine Triphosphate
Proteins

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

Global Effects of DDX3 Inhibition on Cell Cycle Regulation Identified by a Combined Phosphoproteomics and Single Cell Tracking Approach. / Heerma van Voss, Marise R.; Kammers, Kai; Vesuna, Farhad; Brilliant, Justin; Bergman, Yehudit; Tantravedi, Saritha; Wu, Xinyan; Cole, Robert N; Holland, Andrew; van Diest, Paul J.; Raman, Venu.

In: Translational Oncology, Vol. 11, No. 3, 01.06.2018, p. 755-763.

Research output: Contribution to journalArticle

Heerma van Voss, Marise R. ; Kammers, Kai ; Vesuna, Farhad ; Brilliant, Justin ; Bergman, Yehudit ; Tantravedi, Saritha ; Wu, Xinyan ; Cole, Robert N ; Holland, Andrew ; van Diest, Paul J. ; Raman, Venu. / Global Effects of DDX3 Inhibition on Cell Cycle Regulation Identified by a Combined Phosphoproteomics and Single Cell Tracking Approach. In: Translational Oncology. 2018 ; Vol. 11, No. 3. pp. 755-763.
@article{4c6e6e5dc8a346258cdfc4aa13088c3c,
title = "Global Effects of DDX3 Inhibition on Cell Cycle Regulation Identified by a Combined Phosphoproteomics and Single Cell Tracking Approach",
abstract = "DDX3 is an RNA helicase with oncogenic properties. The small molecule inhibitor RK-33 is designed to fit into the ATP binding cleft of DDX3 and hereby block its activity. RK-33 has shown potent activity in preclinical cancer models. However, the mechanism behind the antineoplastic activity of RK-33 remains largely unknown. In this study we used a dual phosphoproteomic and single cell tracking approach to evaluate the effect of RK-33 on cancer cells. MDA-MB-435 cells were treated for 24 hours with RK-33 or vehicle control. Changes in phosphopeptide abundance were analyzed with quantitative mass spectrometry using isobaric mass tags (Tandem Mass Tags). At the proteome level we mainly observed changes in mitochondrial translation, cell division pathways and proteins related to cell cycle progression. Analysis of the phosphoproteome indicated decreased CDK1 activity after RK-33 treatment. To further evaluate the effect of DDX3 inhibition on cell cycle progression over time, we performed timelapse microscopy of Fluorescent Ubiquitin Cell Cycle Indicators labeled cells after RK-33 or siDDX3 exposure. Single cell tracking indicated that DDX3 inhibition resulted in a global delay in cell cycle progression in interphase and mitosis. In addition, we observed an increase in endoreduplication. Overall, we conclude that DDX3 inhibition affects cells in all phases and causes a global cell cycle progression delay.",
author = "{Heerma van Voss}, {Marise R.} and Kai Kammers and Farhad Vesuna and Justin Brilliant and Yehudit Bergman and Saritha Tantravedi and Xinyan Wu and Cole, {Robert N} and Andrew Holland and {van Diest}, {Paul J.} and Venu Raman",
year = "2018",
month = "6",
day = "1",
doi = "10.1016/j.tranon.2018.04.001",
language = "English (US)",
volume = "11",
pages = "755--763",
journal = "Translational Oncology",
issn = "1936-5233",
publisher = "Neoplasia Press",
number = "3",

}

TY - JOUR

T1 - Global Effects of DDX3 Inhibition on Cell Cycle Regulation Identified by a Combined Phosphoproteomics and Single Cell Tracking Approach

AU - Heerma van Voss, Marise R.

AU - Kammers, Kai

AU - Vesuna, Farhad

AU - Brilliant, Justin

AU - Bergman, Yehudit

AU - Tantravedi, Saritha

AU - Wu, Xinyan

AU - Cole, Robert N

AU - Holland, Andrew

AU - van Diest, Paul J.

AU - Raman, Venu

PY - 2018/6/1

Y1 - 2018/6/1

N2 - DDX3 is an RNA helicase with oncogenic properties. The small molecule inhibitor RK-33 is designed to fit into the ATP binding cleft of DDX3 and hereby block its activity. RK-33 has shown potent activity in preclinical cancer models. However, the mechanism behind the antineoplastic activity of RK-33 remains largely unknown. In this study we used a dual phosphoproteomic and single cell tracking approach to evaluate the effect of RK-33 on cancer cells. MDA-MB-435 cells were treated for 24 hours with RK-33 or vehicle control. Changes in phosphopeptide abundance were analyzed with quantitative mass spectrometry using isobaric mass tags (Tandem Mass Tags). At the proteome level we mainly observed changes in mitochondrial translation, cell division pathways and proteins related to cell cycle progression. Analysis of the phosphoproteome indicated decreased CDK1 activity after RK-33 treatment. To further evaluate the effect of DDX3 inhibition on cell cycle progression over time, we performed timelapse microscopy of Fluorescent Ubiquitin Cell Cycle Indicators labeled cells after RK-33 or siDDX3 exposure. Single cell tracking indicated that DDX3 inhibition resulted in a global delay in cell cycle progression in interphase and mitosis. In addition, we observed an increase in endoreduplication. Overall, we conclude that DDX3 inhibition affects cells in all phases and causes a global cell cycle progression delay.

AB - DDX3 is an RNA helicase with oncogenic properties. The small molecule inhibitor RK-33 is designed to fit into the ATP binding cleft of DDX3 and hereby block its activity. RK-33 has shown potent activity in preclinical cancer models. However, the mechanism behind the antineoplastic activity of RK-33 remains largely unknown. In this study we used a dual phosphoproteomic and single cell tracking approach to evaluate the effect of RK-33 on cancer cells. MDA-MB-435 cells were treated for 24 hours with RK-33 or vehicle control. Changes in phosphopeptide abundance were analyzed with quantitative mass spectrometry using isobaric mass tags (Tandem Mass Tags). At the proteome level we mainly observed changes in mitochondrial translation, cell division pathways and proteins related to cell cycle progression. Analysis of the phosphoproteome indicated decreased CDK1 activity after RK-33 treatment. To further evaluate the effect of DDX3 inhibition on cell cycle progression over time, we performed timelapse microscopy of Fluorescent Ubiquitin Cell Cycle Indicators labeled cells after RK-33 or siDDX3 exposure. Single cell tracking indicated that DDX3 inhibition resulted in a global delay in cell cycle progression in interphase and mitosis. In addition, we observed an increase in endoreduplication. Overall, we conclude that DDX3 inhibition affects cells in all phases and causes a global cell cycle progression delay.

UR - http://www.scopus.com/inward/record.url?scp=85045621039&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85045621039&partnerID=8YFLogxK

U2 - 10.1016/j.tranon.2018.04.001

DO - 10.1016/j.tranon.2018.04.001

M3 - Article

C2 - 29684792

AN - SCOPUS:85045621039

VL - 11

SP - 755

EP - 763

JO - Translational Oncology

JF - Translational Oncology

SN - 1936-5233

IS - 3

ER -