Predicted secondary structures and sequence comparisons of EFhd2 and EFhd1. a) Predicted secondary structures of murine EFhd1 and EFhd2 (240 aa) (http://elm.eu.org). PR: proline-rich elements (potential SH3 domain-binding sites), EF: EF-hands, CC: coiled-coil domain, D: disordered region.

b) Aligment of murine EFhd1 (Q9D4J1) and EFhd2 (Q9D8Y0) sequences using ClustalW2 (http://www.ebi.ac.uk/clustalw). Amino acid (aa) positions are marked on the right. "*", identical aa, ":", conserved aa, "." semi conserved aa.

Phylogeny of EFhd genes. A phylogenetic tree with equal distances between different taxons was generated using efhd1 as search parameter (http://www.treefam.org/cgi-bin). Search parameters were: accession number TF320736 / clean / equal distance / show taxa / none / exp / selected species. Only representative species are shown for convenience. On the right hand side the employed transcript sequences are shown. Chick, chicken (Gallus gallus), Brare, zebrafish (Danio rerio), Xentr, frog (Xenopus tropicalis), Cioin, Hydra (Ciona intestinalis), Drome, fruitfly (Drosophila melanogaster), Caeel, worm (Caenorhabtidis elegans).

Involvement of EFhd2 in membrane and cytoskeleton associated pathways. EFhd2 can associate with detergent resistant membranes (DRM) thought to represent membrane rafts in some B cell lines. Membrane raft association of EFhd2 was also detected in nervous tissue of mice that express a gain of function mutant (G93A) of SOD1. EFhd2 has been shown to exhibit NF-κB-activating as well as NF-κB-inactivating functions in different cell types. Through regulation of NF-κB, EFhd2 may regulate survival or apoptosis of diverse cell types. The interactions of EFhd2 with the SH3 domains of Lyn, PLCγ and Fgr are based on GST-Pull down experiments using recombinant SH3 domains (see Ref. 14). The interactions of Fgr and Lyn SH3 domains with EFhd2 depend on phosphorylation and the SH3 domain of Fgr binds to the predicted proline-rich (PR) SH3 domain binding region of EFhd2. Three phosphorylation sites of EFhd2 are known: S74, S76 and Y83. S74 is a cyclin dependent kinase 1 (Cdk1) site. The ability of EFhd2 to bind calcium may regulate its function and its association with the cytoskeleton. EFhd2 also interacts with hyperphosphorylated Tau in a FTDP17 mouse model as well as in FTDP17 and Alzheimer's patients.

Involvement of EFhd1 in survival and redox regulated pathways. EFhd1 has been described as a survival factor for neuronal cells and been proposed to mediate muscle cell survival. In response to oxidative stress that can be blocked by superoxide dismutase 2 (SOD2), HNF-4α (hepatocyte nuclear factor 4α) becomes activated. EFhd1 is a direct target of HNF-4α and its expression is repressed by SOD2. Thus, EFhd1 may be involved in a protective cellular response against oxidative stress and, thereby, exhibit anti-apoptotic functions in diverse cell types. Early B cell factor 1 (Ebf1) positively regulates EFhd1 expression in early B cells whereas Suz12 as part of the polycomb repressor complex 2 (PRC2) is a negative regulator of EFhd1 expression.