Deep and quantitative profiling of PTMs in ecologically preserved and polluted pollen proteomes of timothy grass reveals predominant source of contamination

Abstract

Objective: to create method for unrestrictive deep, relative quantification of post translationalmodifications (PTMs) within different proteomes. Pollution field studies of bio indicators such aspollen are valuable because of realistic situation of target contamination, however they carry thegreat extent of uncertainty in attributing and delineating the polluting effect from multiple sources.Holistic research platform focusing on comprehensively characterized and quantified PTMs ofcomparable bio-indicator proteomes could help and overcome these obstacles of field pollutionstudies.Material and Methods: Scanning electron and light microscopy assessed surface and sub pollenparticle (SPP) releasing features of timothy grass (TG) pollen. Inductively coupled atomic emissionspectrometry revealed metal elemental content in pollen while in solution trypsin digested pollenproteomes analysed with high resolution Orbitrap mass tandem spectrometry and PEAKS Suite 8.5brought quantitative information on protein expression level and its PTM profiling.Results: TG polluted pollen samples (P2) collected along regional road and chemical plant,exposed to air contaminants from road traffics and chemical plants showed 4.5 times higher SPPreleasing capacity, with notable surface changes, as well as significantly higher contents of Mn, Hgand Cd. Antioxidative enzymes (oxidoreductases, superoxide dismutases and peroxidases),including actin, were upregulated several times in polluted sample compared to ecologicallypreserved pollen (P1). While the level of spontaneous and physiological PTMs includingmethylation, acetylation, deamidation and formylation, was similar without significant changes inP1 and P2 pollens, oxidative PTMs including oxidation of Met, Lys, His, Pro and HNE and hexoseadducts showed several times higher and significant increase in abundancy of P2 compared to P1.PTMs connected to road traffic such as tyrosine nitration were very rare and low abundant.Conclusion: Results suggest prominent role of chemical pollution compared to effect of road trafficpollution, with primary consequences from oxidative properties of mercury (Hg) and cadmium(Cd).