Methods

A method for identifying a compound expected to be useful in modulating, for example inhibiting, the phosphorylation of SPAK or OSR1 in a cell, the method comprising the steps of (1) determining whether a test compound modulates, for example inhibits, the protein kinase activity of a WNK isoform polypeptide, and (2) selecting a compound which modulates, for example inhibits, the protein kinase activity of the WNK isoform polypeptide.

A method of identifying a compound expected to be useful in modulating, for example inhibiting, the phosphorylation of SPAK or OSR1 in a cell, comprising the step of determining the effect of the compound on the ability of a WNK isoform polypeptide to bind to a SPAK polypeptide or OSR1 polypeptide and (2) selecting a compound which modulates, for example inhibits, the ability of said WNK isoform polypeptide to bind to the SPAK or OSR1 polypeptide.

A mutated SPAK polypeptide wherein the residue corresponding to Ser373 is mutated, for example to an alanine residue or to an aspartate or glutamate residue;

• and/or wherein the residue corresponding to D212 is mutated, for example to an alanine residue.

A mutated OSR1 polypeptide wherein the T-loop threonine residue corresponding to Thr185 of human OSR1 is mutated, for example to an alanine residue or to an aspartate or glutamate residue;

• and/or wherein the residue corresponding to Ser325 is mutated, for example to an alanine residue or to an aspartate or glutamate residue;
• and/or wherein the residue corresponding to D164 is mutated, for example to an alanine residue.

A method of activating a SPAK polypeptide or OSR1 polypeptide wherein the SPAK polypeptide or OSR1 polypeptide is phosphorylated by a WNK isoform polypeptide.

Use of a WNK isoform polypeptide in a method of phosphorylating a SPAK polypeptide or OSR1 polypeptide.

A specific binding partner, typically an antibody, that binds in a phosphorylation state-sensitive manner to an epitope encompassing Ser373 of SPAK (for example human SPAK), or Thr185 or Ser 325 of OSR1 (for example human OSR1).

A peptide comprising a fragment derivable from SPAK which encompasses the Ser373 residue of SPAK and at least part of the sequence which includes this residue, for example at least the 2, 3, 4, 5, 6 or 7 residues C-terminal or N-terminal of this residue, wherein the fragment has less than 50, 40, 30, 25, 20, 15 or 10 residues.

A peptide comprising a fragment derivable from OSR1 which encompasses the Ser325 residue of OSR1 or the Thr185 residue of OSR1 and at least part of the sequence which includes this residue, for example at least the 2, 3, 4, 5, 6 or 7 residues C-terminal or N-terminal of this residue, wherein the fragment has less than 50, 40, 30, 25, 20, 15 or 10 residues.