Increased abundance of the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1) in patients with obesity and type 2 diabetes: Evidence for altered adiponectin signalling

R. M. Holmes, Z. Yi, E. De Filippis, R. Berria, S. Shahani, P. Sathyanarayana, V. Sherman, K. Fujiwara, C. Meyer, C. Christ-Roberts, H. Hwang, J. Finlayson, L. Q. Dong, L. J. Mandarino, M. Bajaj

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

Aims/hypothesis: The adiponectin signalling pathway is largely unknown, but recently the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1), has been shown to interact directly with adiponectin receptor (ADIPOR)1. APPL1 is present in C2C12 myoblasts and mouse skeletal muscle, but its presence in human skeletal muscle has not been investigated. Methods: Samples from type 2 diabetic, and lean and non-diabetic obese participants were analysed by: immunoprecipitation and western blot; HPLC-electrospray ionisation (ESI)-mass spectrometry (MS) analysis; peak area analysis by MS; HPLC-ESI-MS/MS/MS analysis; and RT-PCR analysis of APPL1 mRNA. Results: Immunoprecipitation and western blot indicated a band specific to APPL1. Tryptic digestion and HPLC-ESI-MS analysis of whole-muscle homogenate APPL1 unambiguously identified APPL1 with 56% sequence coverage. Peak area analysis by MS validated western blot results, showing APPL1 levels to be significantly increased in type 2 diabetic and obese as compared with lean participants. Targeted phosphopeptide analysis by HPLC-ESI-MS/MS/MS showed that APPL1 was phosphorylated specifically on Ser401. APPL1 mRNA expression was significantly increased in obese and type 2 diabetic participants as compared with lean participants. After bariatric surgery in morbidly obese participants with subsequent weight loss, skeletal muscle APPL1 abundance was significantly reduced (p<0.05) in association with an increase in plasma adiponectin (p<0.01), increased levels of ADIPOR1 (p<0.05) and increased muscle AMP-activated protein kinase (AMPK) phosphorylation (p<0.05). Conclusions/interpretation: APPL1 abundance is significantly higher in type 2 diabetic muscle; APPL1 is phosphorylated in vivo on Ser 401. Improvements in hyperglycaemia and hypoadiponectinaemia following weight loss are associated with reduced skeletal muscle APPL1, and increased plasma adiponectin levels and muscle AMPK phosphorylation.

Original languageEnglish (US)
Pages (from-to)2122-2131
Number of pages10
JournalDiabetologia
Volume54
Issue number8
DOIs
StatePublished - Aug 2011
Externally publishedYes

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Leucine Zippers
Phosphotyrosine
Adiponectin
Tandem Mass Spectrometry
Type 2 Diabetes Mellitus
Electrospray Ionization Mass Spectrometry
Obesity
Skeletal Muscle
High Pressure Liquid Chromatography
Muscles
Proteins
AMP-Activated Protein Kinases
Western Blotting
Immunoprecipitation
Weight Loss
Mass Spectrometry
Adiponectin Receptors
Phosphorylation
Phosphopeptides
Messenger RNA

Keywords

  • Adiponectin
  • APPL1
  • Obesity
  • Type 2 diabetes

ASJC Scopus subject areas

  • Internal Medicine
  • Endocrinology, Diabetes and Metabolism

Cite this

Increased abundance of the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1) in patients with obesity and type 2 diabetes : Evidence for altered adiponectin signalling. / Holmes, R. M.; Yi, Z.; De Filippis, E.; Berria, R.; Shahani, S.; Sathyanarayana, P.; Sherman, V.; Fujiwara, K.; Meyer, C.; Christ-Roberts, C.; Hwang, H.; Finlayson, J.; Dong, L. Q.; Mandarino, L. J.; Bajaj, M.

In: Diabetologia, Vol. 54, No. 8, 08.2011, p. 2122-2131.

Research output: Contribution to journalArticle

Holmes, RM, Yi, Z, De Filippis, E, Berria, R, Shahani, S, Sathyanarayana, P, Sherman, V, Fujiwara, K, Meyer, C, Christ-Roberts, C, Hwang, H, Finlayson, J, Dong, LQ, Mandarino, LJ & Bajaj, M 2011, 'Increased abundance of the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1) in patients with obesity and type 2 diabetes: Evidence for altered adiponectin signalling', Diabetologia, vol. 54, no. 8, pp. 2122-2131. https://doi.org/10.1007/s00125-011-2173-x
Holmes, R. M. ; Yi, Z. ; De Filippis, E. ; Berria, R. ; Shahani, S. ; Sathyanarayana, P. ; Sherman, V. ; Fujiwara, K. ; Meyer, C. ; Christ-Roberts, C. ; Hwang, H. ; Finlayson, J. ; Dong, L. Q. ; Mandarino, L. J. ; Bajaj, M. / Increased abundance of the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1) in patients with obesity and type 2 diabetes : Evidence for altered adiponectin signalling. In: Diabetologia. 2011 ; Vol. 54, No. 8. pp. 2122-2131.
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T1 - Increased abundance of the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1) in patients with obesity and type 2 diabetes

T2 - Evidence for altered adiponectin signalling

AU - Holmes, R. M.

AU - Yi, Z.

AU - De Filippis, E.

AU - Berria, R.

AU - Shahani, S.

AU - Sathyanarayana, P.

AU - Sherman, V.

AU - Fujiwara, K.

AU - Meyer, C.

AU - Christ-Roberts, C.

AU - Hwang, H.

AU - Finlayson, J.

AU - Dong, L. Q.

AU - Mandarino, L. J.

AU - Bajaj, M.

PY - 2011/8

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N2 - Aims/hypothesis: The adiponectin signalling pathway is largely unknown, but recently the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1), has been shown to interact directly with adiponectin receptor (ADIPOR)1. APPL1 is present in C2C12 myoblasts and mouse skeletal muscle, but its presence in human skeletal muscle has not been investigated. Methods: Samples from type 2 diabetic, and lean and non-diabetic obese participants were analysed by: immunoprecipitation and western blot; HPLC-electrospray ionisation (ESI)-mass spectrometry (MS) analysis; peak area analysis by MS; HPLC-ESI-MS/MS/MS analysis; and RT-PCR analysis of APPL1 mRNA. Results: Immunoprecipitation and western blot indicated a band specific to APPL1. Tryptic digestion and HPLC-ESI-MS analysis of whole-muscle homogenate APPL1 unambiguously identified APPL1 with 56% sequence coverage. Peak area analysis by MS validated western blot results, showing APPL1 levels to be significantly increased in type 2 diabetic and obese as compared with lean participants. Targeted phosphopeptide analysis by HPLC-ESI-MS/MS/MS showed that APPL1 was phosphorylated specifically on Ser401. APPL1 mRNA expression was significantly increased in obese and type 2 diabetic participants as compared with lean participants. After bariatric surgery in morbidly obese participants with subsequent weight loss, skeletal muscle APPL1 abundance was significantly reduced (p<0.05) in association with an increase in plasma adiponectin (p<0.01), increased levels of ADIPOR1 (p<0.05) and increased muscle AMP-activated protein kinase (AMPK) phosphorylation (p<0.05). Conclusions/interpretation: APPL1 abundance is significantly higher in type 2 diabetic muscle; APPL1 is phosphorylated in vivo on Ser 401. Improvements in hyperglycaemia and hypoadiponectinaemia following weight loss are associated with reduced skeletal muscle APPL1, and increased plasma adiponectin levels and muscle AMPK phosphorylation.

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KW - Adiponectin

KW - APPL1

KW - Obesity

KW - Type 2 diabetes

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