Zebra fish aquaporins
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Zebra Fish Aquaporins. Magdalena Calusinska Ang è le Tingaud – Sequeira David Otero Joan Cerd à. 1991 – discovery of water channels. 2003 – Nobel Prize in Chemistry for Peter Agre for the discovery of aquaporins. Water – natural environment of fish.

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Zebra fish aquaporins

Zebra Fish Aquaporins

Magdalena Calusinska

Angèle Tingaud – Sequeira

David Otero

Joan Cerdà

1991 discovery of water channels
1991 – discovery of water channels

2003 – Nobel Prize in Chemistry for Peter Agre for the discovery of aquaporins

Major intrinsic proteins mips aquaporins
Major intrinsic proteins (MIPs) – Aquaporins

  • homotetramers consisting of four monomeric channels

  • composed of single polypeptide chain ~ 270aa

  • six transmembrane α helices, with three extracellular loops (A,C,E) and two intracellular (B,D)

  • N and C terminal ends reside in the cytoplasm

  • highly conserved NPA motifs are present in loop B and E

de Groot and Grubmüller, 2005

Water and glycerol transport
Water and glycerol transport

de Groot and Grubmüller, 2005

Aquaporins in metazoa
Aquaporins in Metazoa

  • 13 paralogs have been identified so far in mammals

  • they are present in many different cells and tissues

  • they have a unique cellular and subcellular distribution with little overlap between homologs

  • in mammals amino acid sequences of paralogs show 20 – 60% identity

  • they differ mainly at the intracellular N – and C – termini (<20%)

Heymann and Engel, 1999

Human aquaporin family gene cluster
Human aquaporin family gene cluster

Organization map of an aquaporin cluster in human

AQP2, 5 and 6 are absent from

the genome of Danio rerio

Distance between AQP0 and the AQP2, 5 and 6

gene cluster is approximately 500kb

Dr AQP0a, Dr AQP0b

Zebra Fish chromosome 23.

No sequences corresponding to

AQP2, 5 and 6 could be mapped

Aquaporins in zebra fish
Aquaporins in Zebra Fish



Neighbour – joining tree based on the

amino acid, full coding sequences of

Zebra Fish AQPs

Aquaporins and aquaglyceroporins
Aquaporins and Aquaglyceroporins

Aquaglyceroporins contain two

Additional peptide spans located in

Loop C and E respectively

Permeability properties xenopus laevis oocyte swelling assay
Permeability properties – Xenopus laevisoocyte swelling assay

Injection of cRNA

Surgical recovery of

stage V and VI oocytes

Xenopus laevis

  • Swelling measurement:

  • 200mOsm → 20mOsm

  • 12 pictures every 2s

  • calculation of Pf

Water permeability p f hg 2 sensitivity
Water permeability – Pf / Hg2+ sensitivity


Conditions used: 1 ng of cRNA injetced

0,5 mM Hg2+


Dr aqp3b isoform
Dr AQP3b isoform

Similarity to mammaliam AQP6??

Dr AQP3a doesn’t show sensitivity

to Hg ions. Contrary, the water transport is stimulated after incubation with 0,5 mM Hg2+.

Dr aqp3b a possible ion channel
Dr AQP3b – a possible ion channel?

Unique residues implicated

in AQP6 ion conductance

Alignment of mammalian

AQP6 and Dr AQP3b

Structural model of highlighting

the crossing point between

TM2 and TM5

Liu et al., 2005

Heterotetrameric composition of dr aqp4
Heterotetrameric composition of Dr AQP4??



*:. .* : :* ..** : ***.******** **:**:.*****:******:******

1 ng of cRNA injected


Neely et al., 1999

* p=0,0062

Glycerol permeability p gly
Glycerol permeability - Pgly


Dr AQP1a used

as a negative control

Aquaglyceroporins used in this study:

Dr AQP1a – 1ng cRNA injected

Dr AQP3b – 10ng cRNA injected

Dr AQP9b – 1ng cRNA injected

Dr AQP10a – 1ng cRNA injected


  • 9 orthologs of AQPs out of 13 existing in mammals are present in Zebra Fish

  • Dr AQPs 0,1,9 represent two isoforms and Dr AQPs 8 and 10 three

  • No homologies to mammalian AQPs 2,5 and 6 have been found in fish

Future studies on zebra fish aqps
Future studies on Zebra Fish AQPs

  • Expression pattern of different AQPs isoforms in Zebra Fish tissues

  • Comparison of efficiency of water transport between different Dr AQPs – tagged proteins

  • ISH of AQPs in embryos

  • Permeability of selected AQPs to CPAs