Cytoskeletal and motor proteins facilitate trafficking of AQP1-containing vesicles in cholangiocytes

Pamela S. Tietz, Mark A Mc Niven, Patrick L. Splinter, Bing Q. Huang, Nicholas F La Russo

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

Background information. We have previously showed that: (i) cholangiocytes contain AQP1 (aquaporin 1) water channels sequestered in intracellular vesicles; and (ii) upon stimulation with choleretic agonists such as secretin or dibutyryl-cAMP (dbcAMP), the AQP1 vesicles move via microtubules to the apical cholangiocyte membrane to facilitate osmotically driven, passive water movement (i.e. ductal bile secretion). The aim of the present study was to determine which proteins and mechanisms regulate AQP1 trafficking in cholangiocytes. Results. Using polarized cultured NMCs (normal mouse cholangiocytes) or NRCs (normal rat cholangiocytes) and affinity-purified antibodies, we performed immunofluorescent confocal microscopy on fixed cells or immunoblotting on cell lysates for actin, tubulin, kinesin and dynein, proteins known to regulate intracellular vesicle trafficking. By immunostaining, the appropriate orientation of the actin (i.e. sub-apical) and tubulin (i.e. generalized) cytoskeleton was apparent; kinesin and dynein displayed a homogeneous punctate distribution. Immunoblotting showed kinesin and dynein to be present in both cholangiocyte lysates and in isolated AQP1-containing vesicles. We utilized real-time fluorescence confocal microscopy of NMCs transfected with a GFP (green fluorescent protein)-AQP1 fusion construct in the presence and absence of dbcAMP. Conclusions. Our results provide additional insights into the potential molecular mechanisms of ductal bile secretion.

Original languageEnglish (US)
Pages (from-to)43-52
Number of pages10
JournalBiology of the Cell
Volume98
Issue number1
DOIs
StatePublished - Jan 2006

Fingerprint

Aquaporin 1
Cytoskeletal Proteins
Protein Transport
Dyneins
Kinesin
Tubulin
Immunoblotting
Confocal Microscopy
Bile
Actins
Cholagogues and Choleretics
Water Movements
Aquaporins
Antibody Affinity
Secretin
Green Fluorescent Proteins
Cytoskeleton
Fluorescence Microscopy
Microtubules
Proteins

Keywords

  • Aquaporin-1
  • Cholangiocyte
  • Dynein
  • Kinesin
  • Vesicle

ASJC Scopus subject areas

  • Cell Biology

Cite this

Cytoskeletal and motor proteins facilitate trafficking of AQP1-containing vesicles in cholangiocytes. / Tietz, Pamela S.; Mc Niven, Mark A; Splinter, Patrick L.; Huang, Bing Q.; La Russo, Nicholas F.

In: Biology of the Cell, Vol. 98, No. 1, 01.2006, p. 43-52.

Research output: Contribution to journalArticle

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