HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) All Versions of this Article: M700596-MCP200v1 7/10/1998    most recent Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Glossary Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Espina, V. Articles by Liotta, L. A. Search for Related Content PubMed PubMed Citation Articles by Espina, V. Articles by Liotta, L. A. Social Bookmarking                      What's this?

Molecular & Cellular Proteomics 7:1998-2018, 2008.
© 2008 by The American Society for Biochemistry and Molecular Biology, Inc.

---

Research

A Portrait of Tissue Phosphoprotein Stability in the Clinical Tissue Procurement Process^*

Virginia Espina^,, Kirsten H. Edmiston^¶, Michael Heiby, Mariaelena Pierobon^,||, Manuela Sciro^,**, Barbara Merritt^¶, Stacey Banks^¶, Jianghong Deng, Amy J. VanMeter, David H. Geho^,, Lucia Pastore, Joel Sennesh, Emanuel F. Petricoin, III and Lance A. Liotta

From the Center for Applied Proteomics and Molecular Medicine, George Mason University, Manassas, Virginia 20110, ^|| Clinica Chirurcica II, Università degli Studi di Padova, 35128 Padova (Padova), Italy, ^** Centro di Referimento Oncologico, National Cancer Institute, Aviano Hospital, 33081 Aviano (Pordenone), Italy, and ^¶ Inova Cancer Center and Department of Pathology, Inova Fairfax Hospital, Falls Church, Virginia 22042

Little is known about the preanalytical fluctuations of phosphoproteins during tissue procurement for molecular profiling. This information is crucial to establish guidelines for the reliable measurement of these analytes. To develop phosphoprotein profiles of tissue subjected to the trauma of excision, we measured the fidelity of 53 signal pathway phosphoproteins over time in tissue specimens procured in a community clinical practice. This information provides strategies for potential surrogate markers of stability and the design of phosphoprotein preservative/fixation solutions. Eleven different specimen collection time course experiments revealed augmentation (±20% from the time 0 sample) of signal pathway phosphoprotein levels as well as decreases over time independent of tissue type, post-translational modification, and protein subcellular location (tissues included breast, colon, lung, ovary, and uterus (endometrium/myometrium) and metastatic melanoma). Comparison across tissue specimens showed an >20% decrease of protein kinase B (AKT) Ser-473 (p < 0.002) and myristoylated alanine-rich C-kinase substrate protein Ser-152/156 (p < 0.0001) within the first 90-min postexcision. Proteins in apoptotic (cleaved caspase-3 Asp-175 (p < 0.001)), proliferation/survival/hypoxia (IRS-1 Ser-612 (p < 0.0003), AMP-activated protein kinase β Ser-108 (p < 0.005), ERK Thr-202/Tyr-204 (p < 0.003), and GSK3β Ser-21/9 (p < 0.01)), and transcription factor pathways (STAT1 Tyr-701 (p < 0.005) and cAMP response element-binding protein Ser-133 (p < 0.01)) showed >20% increases within 90-min postprocurement. Endothelial nitric-oxide synthase Ser-1177 did not change over the time period evaluated with breast or leiomyoma tissue. Treatment with phosphatase or kinase inhibitors alone revealed that tissue kinase pathways are active ex vivo. Combinations of kinase and phosphatase inhibitors appeared to stabilize proteins that exhibited increases in the presence of phosphatase inhibitors alone (ATF-2 Thr-71, SAPK/JNK Thr-183/Tyr-185, STAT1 Tyr-701, JAK1 Tyr-1022/1023, and PAK1/PAK2 Ser-199/204/192/197). This time course study 1) establishes the dynamic nature of specific phosphoproteins in excised tissue, 2) demonstrates augmented phosphorylation in the presence of phosphatase inhibitors, 3) shows that kinase inhibitors block the upsurge in phosphorylation of phosphoproteins, 4) provides a rational strategy for room temperature preservation of proteins, and 5) constitutes a foundation for developing evidence-based tissue procurement guidelines.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				S. Baginsky and W. Gruissem The Chloroplast Kinase Network: New Insights from Large-Scale Phosphoproteome Profiling Mol Plant, November 1, 2009; 2(6): 1141 - 1153. [Abstract] [Full Text] [PDF]

				J. Bodo, L. Durkin, and E. D. Hsi Quantitative In Situ Detection of Phosphoproteins in Fixed Tissues Using Quantum Dot Technology J. Histochem. Cytochem., July 1, 2009; 57(7): 701 - 708. [Abstract] [Full Text] [PDF]

				S. Reiland, G. Messerli, K. Baerenfaller, B. Gerrits, A. Endler, J. Grossmann, W. Gruissem, and S. Baginsky Large-Scale Arabidopsis Phosphoproteome Profiling Reveals Novel Chloroplast Kinase Substrates and Phosphorylation Networks Plant Physiology, June 1, 2009; 150(2): 889 - 903. [Abstract] [Full Text] [PDF]