Colour coded: Bird feather development influenced by protein networks

Feather colour formation

Image: NatureGate

The variation of colour in the plumage of birds is an important factor in their evolutionary development, but little is known about the changes in the underlying proteome that contribute to the changes. So, scientists in Finland from the University of Turku have undertaken a proteomics study of the feather development, using the pied flycatcher, Ficedula hypoleuca, as their model bird.

Erica Leder and colleagues selected this particular bird because breeding males exhibit wide plumage colour variations ranging from dull brown to strong black. It has been established that the colour differences are caused by the relative proportions of two melanin derivatives, eumelanin and phaeomelanin. Moreover, the colour has been shown to affect the behaviour of the species, by enabling species recognition and influencing sexual selection and the aggression of one male towards another.

Feathers were collected from ten young males, five brown and five black birds representing the colour extremes. The proteins were extracted from the quills, closest to the body of the bird, and separated by SDS-PAGE before analysis by tandem mass spectrometry. Their identities were confirmed by searching the MS/MS spectra against the zebra finch database and they were quantified from the normalised spectral abundance factors.

Protein interaction networks

Almost 300 proteins were identified by these classical methods, with 65 of them associated with epidermal growth or pigmentation. The total number of proteins is likely to be higher, since peptide sequences that differed widely between the pied flycatcher and the reference zebra finch would not be identified.

When the protein-protein interaction networks were established from the results, there were some notable differences between the brown and the black phenotypes. One network in particular caught the attention of the researchers, because its expression level was 20% higher in the black phenotype. It was involved with cell-cell signalling, immune cell trafficking and haematological system development.

None of those three processes is directly related to pigmentation but previous work had established a link between immunocompetence and pigmentation in barn owls and tawny oils, so it is not an unknown relationship. In addition, several proteins associated with melanosomes and pigmentation were identified in this network.

A second set of proteins that differed between the two phenotypes were involved with oxidative stress, with free radical scavenging more important in the brown birds. The production of the pigments eumelanin and phaeomelanin are related to increased oxidative stress so mechanisms must be in place to deal with this change. At this stage, the researchers were unable to say whether the changes they observed were definitely due to differences in pigmentation.

The protein network associated with energy balance was also different between the brown and black pigmented birds. This is in agreement with other studies on male pied flycatchers in which the resting metabolic rate was correlated with the darkness of the plumage. In addition, their courting behaviour was affected by the ambient temperature, which could also be related to metabolic differences.

Taken together, the results provide a starting point for a more detailed investigation into the molecular basis behind the variations in plumage colour of the pied flycatcher. The various signalling networks that were exposed might also help to understand the evolutionary significance of the colours of bird feathers.