The Cloning and Characterization of GFP-like Proteins From the Abnormally Pink Pigmented Tissues of Porites lobata
Bridges, Mary Catherine
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Fluorescent proteins (FPs) form a family of proteins with homologies to green fluorescent protein (GFP), originally isolated from the bioluminescent jellyfish <italic>Aequorea victoria<italic>; the majority of this family's members have been cloned from coral species, yielding FPs with emission wavelengths along the entire visible spectrum. In compromised tissues of <italic>Porites<italic> species, a localized visible pink pigmentation exhibiting red fluorescence has been reported; this pathology has been described most recently as an innate immune response. Here we report the cloning, expression, and characterization of a novel red fluorescent protein (tprRFP) and purple chromoprotein (tprCP) from the pink tissue of a <italic>Porites lobata<italic> specimen. The recombinant tprRFP produces a bright red fluorescence with an Em. Max of 614 nm (Ex. Max: 578 nm), similar to the endogenous red fluorescence of the pigmented coral tissue. This 614 nm emission makes tprRFP the furthest red-shifted, naturally occurring protein isolated. This property, as well as its inherent brightness, photostability, and pH stability characterized <italic>in vitro<italic>, make it an excellent candidate for deep tissue and cell imaging applications. The non-fluorescent chromoprotein, tprCP, absorbs maximally at 581 nm, resulting in its visible purple coloration, which is visually distinct from the pink of the <italic>P. lobata<italic> tissue and recombinant tprRFP. We suggest tprRFP is mostly responsible for the visible pink coloration of the coral tissue. The isolation of the tprRFP/CP coding sequences provide the means to more thoroughly investigate the underlying cellular pathology in corals displaying pink tissue pigmentation response as well as evaluate proposed specific roles for these GFP-like proteins in this response.