Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro

Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines. Ausführliche Beschreibung

Gespeichert in:

Autor*in:	Wisam Nabeel Ibrahim [verfasserIn] Abd Almonem Doolaanea [verfasserIn] Mohammad Syaiful Bahari Bin Abdull Rasad [verfasserIn]

Format:	E-Artikel
Sprache:	Englisch

Erschienen:	2018

Schlagwörter:	YB-1 MMPs collagenases shRNA malignant melanoma

Übergeordnetes Werk:	In: Cells - MDPI AG, 2012, 7(2018), 1, p 7
Übergeordnetes Werk:	volume:7 ; year:2018 ; number:1, p 7

Links:	Link aufrufen Link aufrufen Link aufrufen Journal toc

DOI / URN:	10.3390/cells7010007

Katalog-ID:	DOAJ038604124

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allfields	10.3390/cells7010007 doi (DE-627)DOAJ038604124 (DE-599)DOAJ884a4ace73664ba08b3b252a2daad1ad DE-627 ger DE-627 rakwb eng QH301-705.5 Wisam Nabeel Ibrahim verfasserin aut Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro 2018 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines. YB-1 MMPs collagenases shRNA malignant melanoma Biology (General) Abd Almonem Doolaanea verfasserin aut Mohammad Syaiful Bahari Bin Abdull Rasad verfasserin aut In Cells MDPI AG, 2012 7(2018), 1, p 7 (DE-627)718622081 (DE-600)2661518-6 20734409 nnns volume:7 year:2018 number:1, p 7 https://doi.org/10.3390/cells7010007 kostenfrei https://doaj.org/article/884a4ace73664ba08b3b252a2daad1ad kostenfrei http://www.mdpi.com/2073-4409/7/1/7 kostenfrei https://doaj.org/toc/2073-4409 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2018 1, p 7
spelling	10.3390/cells7010007 doi (DE-627)DOAJ038604124 (DE-599)DOAJ884a4ace73664ba08b3b252a2daad1ad DE-627 ger DE-627 rakwb eng QH301-705.5 Wisam Nabeel Ibrahim verfasserin aut Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro 2018 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines. YB-1 MMPs collagenases shRNA malignant melanoma Biology (General) Abd Almonem Doolaanea verfasserin aut Mohammad Syaiful Bahari Bin Abdull Rasad verfasserin aut In Cells MDPI AG, 2012 7(2018), 1, p 7 (DE-627)718622081 (DE-600)2661518-6 20734409 nnns volume:7 year:2018 number:1, p 7 https://doi.org/10.3390/cells7010007 kostenfrei https://doaj.org/article/884a4ace73664ba08b3b252a2daad1ad kostenfrei http://www.mdpi.com/2073-4409/7/1/7 kostenfrei https://doaj.org/toc/2073-4409 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2018 1, p 7
allfields_unstemmed	10.3390/cells7010007 doi (DE-627)DOAJ038604124 (DE-599)DOAJ884a4ace73664ba08b3b252a2daad1ad DE-627 ger DE-627 rakwb eng QH301-705.5 Wisam Nabeel Ibrahim verfasserin aut Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro 2018 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines. YB-1 MMPs collagenases shRNA malignant melanoma Biology (General) Abd Almonem Doolaanea verfasserin aut Mohammad Syaiful Bahari Bin Abdull Rasad verfasserin aut In Cells MDPI AG, 2012 7(2018), 1, p 7 (DE-627)718622081 (DE-600)2661518-6 20734409 nnns volume:7 year:2018 number:1, p 7 https://doi.org/10.3390/cells7010007 kostenfrei https://doaj.org/article/884a4ace73664ba08b3b252a2daad1ad kostenfrei http://www.mdpi.com/2073-4409/7/1/7 kostenfrei https://doaj.org/toc/2073-4409 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2018 1, p 7
allfieldsGer	10.3390/cells7010007 doi (DE-627)DOAJ038604124 (DE-599)DOAJ884a4ace73664ba08b3b252a2daad1ad DE-627 ger DE-627 rakwb eng QH301-705.5 Wisam Nabeel Ibrahim verfasserin aut Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro 2018 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines. YB-1 MMPs collagenases shRNA malignant melanoma Biology (General) Abd Almonem Doolaanea verfasserin aut Mohammad Syaiful Bahari Bin Abdull Rasad verfasserin aut In Cells MDPI AG, 2012 7(2018), 1, p 7 (DE-627)718622081 (DE-600)2661518-6 20734409 nnns volume:7 year:2018 number:1, p 7 https://doi.org/10.3390/cells7010007 kostenfrei https://doaj.org/article/884a4ace73664ba08b3b252a2daad1ad kostenfrei http://www.mdpi.com/2073-4409/7/1/7 kostenfrei https://doaj.org/toc/2073-4409 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2018 1, p 7
allfieldsSound	10.3390/cells7010007 doi (DE-627)DOAJ038604124 (DE-599)DOAJ884a4ace73664ba08b3b252a2daad1ad DE-627 ger DE-627 rakwb eng QH301-705.5 Wisam Nabeel Ibrahim verfasserin aut Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro 2018 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines. YB-1 MMPs collagenases shRNA malignant melanoma Biology (General) Abd Almonem Doolaanea verfasserin aut Mohammad Syaiful Bahari Bin Abdull Rasad verfasserin aut In Cells MDPI AG, 2012 7(2018), 1, p 7 (DE-627)718622081 (DE-600)2661518-6 20734409 nnns volume:7 year:2018 number:1, p 7 https://doi.org/10.3390/cells7010007 kostenfrei https://doaj.org/article/884a4ace73664ba08b3b252a2daad1ad kostenfrei http://www.mdpi.com/2073-4409/7/1/7 kostenfrei https://doaj.org/toc/2073-4409 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 7 2018 1, p 7
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spellingShingle	Wisam Nabeel Ibrahim misc QH301-705.5 misc YB-1 misc MMPs misc collagenases misc shRNA misc malignant melanoma misc Biology (General) Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro
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topic_title	QH301-705.5 Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro YB-1 MMPs collagenases shRNA malignant melanoma
topic	misc QH301-705.5 misc YB-1 misc MMPs misc collagenases misc shRNA misc malignant melanoma misc Biology (General)
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title	Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro
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title_full	Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro
author_sort	Wisam Nabeel Ibrahim
journal	Cells
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author_browse	Wisam Nabeel Ibrahim Abd Almonem Doolaanea Mohammad Syaiful Bahari Bin Abdull Rasad
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format_se	Elektronische Aufsätze
author-letter	Wisam Nabeel Ibrahim
doi_str_mv	10.3390/cells7010007
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title_sort	effect of shrna mediated silencing of yb-1 protein on the expression of matrix collagenases in malignant melanoma cell in vitro
callnumber	QH301-705.5
title_auth	Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro
abstract	Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines.
abstractGer	Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines.
abstract_unstemmed	Background and Objective: YB-1 is a transcription and oncogenic factor capable of binding to DNA and RNA performing versatile functions within normal and cancer cells. Some studies reported the binding of YB-1 with a collagenases gene promoter and influencing their expression. In addition, the role of YB-1 in malignant melanoma was not elucidated. Thus, in this study, the aim was to knock down the expression of YB-1 in A375 malignant melanoma cancer cell using the shRNA approach and study its effect on cancer cell proliferation, migration, and expression of collagenases. Methods: A375 malignant melanoma cell lines were grown in standard conditions and were transfected with three plasmids containing a retroviral pGFP-V-RS vector, two of them containing targeting sequences for YB-1 mRNA. The third plasmid contained a scrambled mRNA sequence as a negative control. Expression of YB-1 was validated using immune-fluorescence staining, RT-PCR and western blotting. The cancer cell proliferation was determined using MTT assay, serial trypan blue cell counting and cell cycle flow-cytometry analysis. Expression of collagenases (MMP1, MMP8, and MMP13) was evaluated using RT-PCR and western blotting analysis. In addition, a wound-healing assay was used to assess cell migration potential. Statistical analysis was performed using one-way ANOVA test with Bonferroni post hoc analysis to compare the quantitative results among samples. Results: The established silenced cell strains (P1 and P2) had nearly 70% knockdown in the expression of YB-1. These YB-1 silenced strains had a significant cell cycle-specific reduction in cell proliferation (p < 0.05 in serial cell counting and cell cycle flow cytometry analysis, p < 0.001 in MTT assay). In addition, YB-1 silenced strains had a remarkable reduction in cell migration potential. Expression of MMP13 was significantly reduced in YB-1 silenced strains. Conclusion: YB-1 oncoprotein is a promising target in the treatment of malignant melanoma. Silencing of this protein is associated with significant anti-proliferative, anti-invasive and MMP13 insulating properties in A375 malignant melanoma cancer cell lines.
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container_issue	1, p 7
title_short	Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro
url	https://doi.org/10.3390/cells7010007 https://doaj.org/article/884a4ace73664ba08b3b252a2daad1ad http://www.mdpi.com/2073-4409/7/1/7 https://doaj.org/toc/2073-4409
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up_date	2024-07-03T18:51:11.314Z
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Effect of shRNA Mediated Silencing of YB-1 Protein on the Expression of Matrix Collagenases in Malignant Melanoma Cell In Vitro

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