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. Author manuscript; available in PMC: 2015 Oct 24.
Published in final edited form as: AFCS Nat Mol Pages. 2011 Jan 18;2011:A001757. doi: 10.1038/mp.a001757.01

Phosphodiesterase 6D, cGMP-specific rod delta

Hannah J Gitschier 1, Rick H Cote 1
PMCID: PMC4618376  NIHMSID: NIHMS729945  PMID: 26504429

Abstract

Network Map States Transitions Functions Protein Classes

Sequence Interactions Pathways Domains & Motifs Protein Structure Orthologs

Sequence Interactions Pathways Domains & Motifs Protein Structure Orthologs Blast Data

Protein Function

In general, the physiological function of prenyl-binding protein (PrBPδ) is poorly understood. Essentially the entire protein comprises a single domain that serves to bind prenyl groups covalently attached to proteins, specifically farnesyl- and geranylgeranyl-containing proteins. Binding of PrBPδ to a prenylated protein serves to solubilize the protein and/or prevent membrane attachment of the prenyl moieties. Thus, the prevailing view is that PrBPδ aids in the transport of prenylated proteins from their site of synthesis to their final destination by sequestering the lipophilic prenyl group during protein transport. The functional significance of PrBPδ binding to non-prenylated proteins is unclear at present.

In mammalian photoreceptor cells (where PrBPδ has been studied most extensively), the current prevailing view is that PrBPδ plays a role in the transport of prenylated proteins from the inner segment to the outer segment where they may function in the photoresponse (Karan etal. 2008). In the retina, PrBPδ has been shown to interact in vitro with the a and β subunits of cyclic nucleotide phosphodiesterase (PDE6), as well as with opsin kinases (G-protein receptor kinase-1 (GRK1) and G-protein receptor kinase-7 (GRK7)) and the retinitis pigmentosa G protein regulator (RPGR) (Florio etal. 1996; Gillespie etal. 1989; Zhang etal. 2005). Further, transport defects are apparent in PrBPδ-deficient mice (Pde6d−/−) as farnesylated GRK1 and geranylgeranylated cone PDE6a subunits are absent from cone outer segment (Zhang etal. 2007).

There is in vitro evidence that PrBPδ can solubilize PDE6 from rod outer segment (ROS) disk membranes and reduce the ability of activated transducin to activate PDE6 (Norton et al. 2005). However, there is currently no support for this regulatory mechanism operating in vivo.

Regulation of Activity

PrBPδ lacks intrinsic enzymatic activity, and is designed to bind prenyl groups within a highly hydrophobic pocket (Hanzal-Bayer etal. 2002). No known regulatory mechanisms have been reported to modulate prenyl binding to PrBPδ.

Binding of PrBPδ to PDE6 does not alter catalytic activity of PDE6, but does enhance nucleotide exchange at the non-catalytic cGMP binding sites of PDE6 (Mou etal. 1999; Gillespie etal. 1989).

Interactions with Ligands and Other Proteins

Ligands that bind to PrBPδ are known as prenyl groups (also referred to as isoprenoids or isoprenyl groups). PrBPδ can interact with farnesyl and geranylgeranyl side chains in the absence of polypeptides but interacts most favorably with prenylated peptides containing methylated C-termini when compared with non-methylated prenylated peptides (Zhang et al. 2004; Zhang etal. 2005; Zhang etal. 2007; Cook etal. 2000).

Numerous prenylated proteins have been shown to interact with PrBPδ in vitro, including PDE6, GRK1, GRK7, Rab8 and Rab13 (Norton etal. 2005; Wilson etal. 1998; Karan etal. 2008; Marzesco etal. 1998; Florio etal. 1996; Gillespie etal. 1989; Zhang etal. 2004; see Zhang etal. 2005 for review).

For the case of PDE6, binding of PrBPδ has been localized by transmission electron microscopy to the catalytic domain of the PDE6 holoenzyme (Goc etal. 2010). Recent evidence suggests that each catalytic subunit of the PDE6 dimer is capable of binding PrBPδ (Florio etal. 1996; Gillespie etal. 1989; Goc etal. 2010).

In addition, PrBPδ interacts in vitro with non-prenylated proteins of the Ras and Rho GTPase family, such as retinitis pigmentosa GTPase regulator (RPGR) (Linari etal. 1999a) and Arl2/Arl3 (Hanzal-Bayer etal. 2002; Linari etal. 1999b), suggesting a possible role for PrBPδ in linking prenylated and non-prenylated proteins (Zhang etal. 2005).

Low-affinity binding of PrBPδ to a PDE5 inhibitor PF-4540124 has been reported as well (Dadvar etal. 2009).

Regulation of Concentration

Quantitative immunoblot assays suggest the stoichiometry of PrBPδ bound to purified, soluble bovine rod PDE6 isolated from bovine retinal extracts is 1.0 +/− 0.1 (Norton etal. 2005). However, the detectable content of PrBPδ in bovine retinal extracts is 0.6 +/− 0.1 mol per mol of PDE6, and in bovine purified ROS is even less, at 0.09 +/− 0.01 mol PrBPδ per mole of PDE6. These data suggest there are sub-stoichiometric concentrations of PrBPδ relative to PDE6 in bovine retina or in photoreceptor cells.

Subcellular Localization

Localization within photoreceptor cells

The subcellular localization of PrBPδ within photoreceptor cells is not entirely clear. In some immunocytochemistry studies in both frog and bovine retina, the majority of PrBPδ immunoreactivity was localized to the inner segment, or the connecting cilium that joins the inner segment and outer segment (Norton et al. 2005). In immunoelectron microscopy studies, PrBPδ was found associated with ciliary axonemes, extending into the rod and cone outer segments. This finding may be indicative of PrBPδ playing a role in prenylated protein transport from the inner segment to the outer segment. However, other immunocytochemistry studies have shown primary localization of PrBPδ to rod outer segment (Zhang etal. 2004; Zhang etal. 2005).

Localization within other cell types

While the subcellular location of endogenous PrBPδ has not been thoroughly studied, a few immunofluorescence experiments indicate that the majority of transfected epitope-tagged PrBPδ is found in the cytosol of endothelial cells with a small portion (~12%) detected in peripheral vesicular compartments along the plasma membrane (Marzesco etal. 1998). Interestingly, deletion of the RVGLFYV C-terminus amino acid sequence in this tagged PrBPδ results in diffusely distributed PrBPδ in the cytoplasm without peripheral staining.

Major Sites of Expression

PrBPδ is ubiquitously expressed in many cell types and tissues, including non-retinal tissues that do not express PDE6 (Florio etal. 1996). PrBPδ mRNA has been detected in many mammalian tissues, including those of brain, heart, adrenal gland, spleen, skeletal muscle, prostate, testis, ovary, small intestine, colon, and peripheral blood, but remains present at the highest level in retinal tissue (Florio etal. 1996). Further, PrBPδ is a highly conserved protein with 99% amino acid sequence identity between human and cow, and 98% amino acid sequence identity between human and mouse. Other homologues have been detected in vertebrates (e.g. chicken, pig, gorilla), invertebrates (e.g. Caenorhabditis elegans with 69% amino acid sequence identity), and even single-celled eukaryotes (e.g. Paramecium) (Lorenz etal. 1998).

Phenotypes

PrBPδ-deficient mice (Pde6d−/−) are fertile and develop normally, but have 20–30% reduced body weight compared with wild-type or heterozygous littermates (Zhang etal. 2007). Additionally, prenylated phototransduction proteins such as rod PDE6, cone PDE6, and GRK1 are downregulated in knockout retinas and mislocalize to the inner segment. Rod and cone physiology is compromised because a slow degeneration of the retina with the shortening of ROS and thinning of the outer nuclear layer is also apparent in PrBPδ-deficient mice (Zhang etal. 2007). Due to the depletion of GRK1 in ROS there is also a delay in the return to the dark-adapted state in rods.

Splice Variants

No splice variants of PrBPδ have been detected to date.

Antibodies

Commercial antibodies are available: mouse monoclonal and rabbit polyclonal (Santa Cruz Biotechnology, several antibodies offered; Affinity BioReagents, #PAI-726).

Full-length (FL) antibody: Rabbit polyclonal antiserum to the PrBPδ protein was raised by immunization with the FL recombinant bovine protein (Norton etal. 2005).

17K-I and 17K-II: synthesized polyclonal antibodies. Both antibodies recognize PrBPδ in immunocytochemistry experiments and immunosorbent assays, but only 17K-II identifies a 17-kDa band in western analysis (Florio etal. 1996).

References

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