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Akt phosphorylation on Thr308 but not on Ser473 correlates with Akt protein kinase activity in human non-small cell lung cancer.

Vincent EE, Elder DJ, Thomas EC, Phillips L, Morgan C, Pawade J, Sohail M, May MT, Hetzel MR, Tavaré JM - Br. J. Cancer (2011)

Bottom Line: Akt activation involves the phosphorylation of two residues: threonine 308 (Thr308) in the activation loop and serine 473 (Ser473) in the C-terminal hydrophobic motif.The phosphorylation status of Thr308 and Ser473 was compared with that of three separate Akt substrates - PRAS40, TSC2 and TBC1D4 - in fresh frozen samples of early-stage human non-small cell lung cancer (NSCLC).Akt Thr308 phosphorylation correlated with the phosphorylation of each Akt substrate tested, whereas Akt Ser473 phosphorylation did not correlate with the phosphorylation of any of the substrates examined.

View Article: PubMed Central - PubMed

Affiliation: School of Biochemistry, Medical Sciences Building, University of Bristol, Bristol BS8 1TD, UK.

ABSTRACT

Background: The activity of the protein kinase Akt is frequently dysregulated in cancer and is an important factor in the growth and survival of tumour cells. Akt activation involves the phosphorylation of two residues: threonine 308 (Thr308) in the activation loop and serine 473 (Ser473) in the C-terminal hydrophobic motif. Phosphorylation of Ser473 has been extensively studied in tumour samples as a correlate for Akt activity, yet the phosphorylation of Thr308 or of downstream Akt substrates is rarely assessed.

Methods: The phosphorylation status of Thr308 and Ser473 was compared with that of three separate Akt substrates - PRAS40, TSC2 and TBC1D4 - in fresh frozen samples of early-stage human non-small cell lung cancer (NSCLC).

Results: Akt Thr308 phosphorylation correlated with the phosphorylation of each Akt substrate tested, whereas Akt Ser473 phosphorylation did not correlate with the phosphorylation of any of the substrates examined.

Conclusion: The phosphorylation of Thr308 is a more reliable biomarker for the protein kinase activity of Akt in tumour samples than Ser473. Any evaluation of the link between Akt phosphorylation or activity in tumour samples and the prediction or prognosis of disease should, therefore, focus on measuring the phosphorylation of Akt on Thr308 and/or at least one downstream Akt substrate, rather than Akt Ser473 phosphorylation alone.

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Related in: MedlinePlus

Examples of Akt Thr308 and Ser473 phosphorylation, together with the phosphorylation of downstream substrates. Triplicate lysates of normal (N1–3) and patient-matched tumour (T1–3) tissues were separated on SDS–PAGE gels. Phosphorylation of Akt on Thr308 and Ser473, and of three downstream substrates, was determined by western blotting with phospho-specific antibodies as indicated. An anti-F1-ATPase antibody was used as a control for protein loading. (A) Examples (patients 11 and 24) of tumours in group Ia. (B) Examples of tumours from two patients (patients 17 and 27) in group Ib. (C) Examples of two patients (patients 4 and 7) in group II.
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fig4: Examples of Akt Thr308 and Ser473 phosphorylation, together with the phosphorylation of downstream substrates. Triplicate lysates of normal (N1–3) and patient-matched tumour (T1–3) tissues were separated on SDS–PAGE gels. Phosphorylation of Akt on Thr308 and Ser473, and of three downstream substrates, was determined by western blotting with phospho-specific antibodies as indicated. An anti-F1-ATPase antibody was used as a control for protein loading. (A) Examples (patients 11 and 24) of tumours in group Ia. (B) Examples of tumours from two patients (patients 17 and 27) in group Ib. (C) Examples of two patients (patients 4 and 7) in group II.

Mentions: The data for the phosphorylation of all three substrates are provided in Supplementary Figures 1–3, and are summarised in the chart of Figure 3. Examples of some of the originating western blot data are provided in Figure 4.


Akt phosphorylation on Thr308 but not on Ser473 correlates with Akt protein kinase activity in human non-small cell lung cancer.

Vincent EE, Elder DJ, Thomas EC, Phillips L, Morgan C, Pawade J, Sohail M, May MT, Hetzel MR, Tavaré JM - Br. J. Cancer (2011)

Examples of Akt Thr308 and Ser473 phosphorylation, together with the phosphorylation of downstream substrates. Triplicate lysates of normal (N1–3) and patient-matched tumour (T1–3) tissues were separated on SDS–PAGE gels. Phosphorylation of Akt on Thr308 and Ser473, and of three downstream substrates, was determined by western blotting with phospho-specific antibodies as indicated. An anti-F1-ATPase antibody was used as a control for protein loading. (A) Examples (patients 11 and 24) of tumours in group Ia. (B) Examples of tumours from two patients (patients 17 and 27) in group Ib. (C) Examples of two patients (patients 4 and 7) in group II.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3111153&req=5

fig4: Examples of Akt Thr308 and Ser473 phosphorylation, together with the phosphorylation of downstream substrates. Triplicate lysates of normal (N1–3) and patient-matched tumour (T1–3) tissues were separated on SDS–PAGE gels. Phosphorylation of Akt on Thr308 and Ser473, and of three downstream substrates, was determined by western blotting with phospho-specific antibodies as indicated. An anti-F1-ATPase antibody was used as a control for protein loading. (A) Examples (patients 11 and 24) of tumours in group Ia. (B) Examples of tumours from two patients (patients 17 and 27) in group Ib. (C) Examples of two patients (patients 4 and 7) in group II.
Mentions: The data for the phosphorylation of all three substrates are provided in Supplementary Figures 1–3, and are summarised in the chart of Figure 3. Examples of some of the originating western blot data are provided in Figure 4.

Bottom Line: Akt activation involves the phosphorylation of two residues: threonine 308 (Thr308) in the activation loop and serine 473 (Ser473) in the C-terminal hydrophobic motif.The phosphorylation status of Thr308 and Ser473 was compared with that of three separate Akt substrates - PRAS40, TSC2 and TBC1D4 - in fresh frozen samples of early-stage human non-small cell lung cancer (NSCLC).Akt Thr308 phosphorylation correlated with the phosphorylation of each Akt substrate tested, whereas Akt Ser473 phosphorylation did not correlate with the phosphorylation of any of the substrates examined.

View Article: PubMed Central - PubMed

Affiliation: School of Biochemistry, Medical Sciences Building, University of Bristol, Bristol BS8 1TD, UK.

ABSTRACT

Background: The activity of the protein kinase Akt is frequently dysregulated in cancer and is an important factor in the growth and survival of tumour cells. Akt activation involves the phosphorylation of two residues: threonine 308 (Thr308) in the activation loop and serine 473 (Ser473) in the C-terminal hydrophobic motif. Phosphorylation of Ser473 has been extensively studied in tumour samples as a correlate for Akt activity, yet the phosphorylation of Thr308 or of downstream Akt substrates is rarely assessed.

Methods: The phosphorylation status of Thr308 and Ser473 was compared with that of three separate Akt substrates - PRAS40, TSC2 and TBC1D4 - in fresh frozen samples of early-stage human non-small cell lung cancer (NSCLC).

Results: Akt Thr308 phosphorylation correlated with the phosphorylation of each Akt substrate tested, whereas Akt Ser473 phosphorylation did not correlate with the phosphorylation of any of the substrates examined.

Conclusion: The phosphorylation of Thr308 is a more reliable biomarker for the protein kinase activity of Akt in tumour samples than Ser473. Any evaluation of the link between Akt phosphorylation or activity in tumour samples and the prediction or prognosis of disease should, therefore, focus on measuring the phosphorylation of Akt on Thr308 and/or at least one downstream Akt substrate, rather than Akt Ser473 phosphorylation alone.

Show MeSH
Related in: MedlinePlus