Expansion for theBrachylophosaurus canadensisCollagen I Sequence and Additional Evidence of the Preservation of Cretaceous Protein

Abstract

Sequence data from biomolecules such as DNA and proteins, which provide critical information for evolutionary studies, have been assumed to be forever outside the reach of dinosaur paleontology. Proteins, which are predicted to have greater longevity than DNA, have been recovered from two nonavian dinosaurs, but these results remain controversial. For proteomic data derived from extinct Mesozoic organisms to reach their greatest potential for investigating questions of phylogeny and paleobiology, it must be shown that peptide sequences can be reliably and reproducibly obtained from fossils and that fragmentary sequences for ancient proteins can be increasingly expanded. To test the hypothesis that peptides can be repeatedly detected and validated from fossil tissues many millions of years old, we applied updated extraction methodology, high-resolution mass spectrometry, and bioinformatics analyses on a Brachylophosaurus canadensis specimen (MOR 2598) from which collagen I peptides were recovered in 2009. We recovered eight peptide sequences of collagen I: two identical to peptides recovered in 2009 and six new peptides. Phylogenetic analyses place the recovered sequences within basal archosauria. When only the new sequences are considered, B. canadensis is grouped more closely to crocodylians, but when all sequences (current and those reported in 2009) are analyzed, B. canadensis is placed more closely to basal birds. The data robustly support the hypothesis of an endogenous origin for these peptides, confirm the idea that peptides can survive in specimens tens of millions of years old, and bolster the validity of the 2009 study. Furthermore, the new data expand the coverage of B. canadensis collagen I (a 33.6% increase in collagen I alpha 1 and 116.7% in alpha 2). Finally, this study demonstrates the importance of reexamining previously studied specimens with updated methods and instrumentation, as we obtained roughly the same amount of sequence data as the previous study with substantially less sample material. Data are available via ProteomeXchange with identifier PXD005087.

Cellular preservation of musculoskeletal specializations in the Cretaceous bird Confuciusornis

“…Further chemical analyses reveal that even some of the original molecular residues of these soft tissues may remain, such as fragments of amino acids from collagen, particularly in the fibrocartilage. This concurs with accruing evidence that some biomolecules may survive, under exceptional circumstances, over many millions of years56789101112. Our reconstruction of the soft tissues around the ankle joint in this Confuciusornis specimen leads to the conclusion that Confuciusornis had evolved a more derived form and, presumably, function of the ankle region (for example, neomorphic, rudimentary tibial cartilage and hypotarsus), revealing new details of how early birds began to adopt more crouched hindlimb postures.…”

Life Inside a Dinosaur Bone: a Thriving Microbiome

“…We detected no evidence of endogenous proteins in the bone studied here and are therefore unable to replicate the claims of protein survival deep into the fossil record, such as the Mesozoic (Pawlicki et al 1966;Schweitzer et al 2005aSchweitzer et al , 2007aSchweitzer et al , 2007bSchweitzer et al , 2008Schweitzer et al , 2009Schweitzer et al , 2013Schweitzer et al , 2014Schweitzer et al , 2016Asara et al 2007;Organ et al 2008;Schweitzer 2011;Bertazzo et al 2015;Cleland et al 2015;Schroeter et al 2017). In contrast, recent Pleistocene-Holocene material can exhibit clear evidence along multiple lines of investigation for endogenous, ancient collagen, even when the fossil (dentine/enamel in this case) is stained black, indicating taphonomic alteration, and the sample is found exhumed in a warm climate and not treated with aseptic techniques.…”

“…Dinosaur bone has been reported to contain endogenous organics such as collagen, osteocytes, and blood vessels. These reports, if verified, could radically change the study of macroevolution and the physiology of extinct organisms given the immense potential of protein sequence data to shed light on the biology and systematics of extinct organisms (Pawlicki et al 1966;Schweitzer et al 2005aSchweitzer et al , 2005bSchweitzer et al , 2007aSchweitzer et al , 2007bSchweitzer et al , 2008Schweitzer et al , 2009Schweitzer et al , 2013Schweitzer et al , 2014Schweitzer et al , 2016Asara et al 2007;Organ et al 2008;Schweitzer 2011;Bertazzo et al 2015;Cleland et al 2015;Schroeter et al 2017). Most of these reports rely on structural observations, mass spectrometry, and immunohistochemistry.…”

Ambiguity coding allows accurate inference of evolutionary parameters from alignments in an aggregated state-space

“…A more speculative and potentially intriguing application would be estimating selection or structural information from ancient protein sequences. Proteins can persist for longer in the environment than DNA under certain conditions [36][37][38], enabling phylogenetic inferences to be made based on substantially older specimens such as dinosaurs [39,40]. Our methods permit the use of a mixture of all available DNA and protein sequences to maximise signal, extending analyses that are normally only possible with DNA sequences to incorporate additional data sources.…”