Copy Number Analysis of the Murine Platelet Proteome Spanning the Complete Abundance Range

Abstract

Knowledge of the identity and quantity of expressed proteins of a cell type is a prerequisite for a complete understanding of its molecular functions. Mass-spectrometry-based proteomics has allowed the identification of the entire protein complement of yeast and the close-to-complete set of proteins expressed in mammalian cell lines. Using recent technological advances, we here characterized the proteome of murine platelets, key actors in mediating hemostasis and thrombosis. We accurately measured the absolute protein concentrations of 13 platelet proteins using SILAC-protein epitope signature tags and used them as reference points to estimate the copy numbers of all proteins of the platelet proteome. To distinguish contaminants such as plasma or erythrocyte proteins from true platelet proteins, we monitored protein abundance profiles across multiple purification steps. In total, we absolutely quantified 4,400 platelet proteins, with estimated copy numbers ranging from less than 10 to about a million per cell. Stoichiometries derived from our data correspond well with previous studies. Our study provides a close-to-complete reference map of platelet proteins that will be useful to the community, for instance, for interpreting mouse models of human platelet diseases.

Coated platelets function in platelet-dependent fibrin formation via integrin α IIb β 3 and transglutaminase factor XIII

“…37,38 Interestingly, in mouse platelets, tissue type transglutaminase (Tgm2) could be detected, albeit with a 25-fold lower expression in comparison to factor XIII (F13a). 28 Our present findings point to a predominant role of factor XIII, and not of tissue type transglutaminase, in both the human and murine systems, which is in line with the recent studies. A role for platelet-derived FXIII was recently proposed in regulating anti-fibrinolysis by crosslinking α 2 -antiplasmin to fibrin.…”

“…These data suggest no major role for tissue type transglutaminase in thrombus formation and Rhod-A14 binding in the absence of coagulation, which is in line with the low expression of tissue-type transglutaminase in mouse platelets. 28 Under coagulant conditions, large fibrin-containing thrombi were formed with both F13a1 +/+ and F13a1 -/-blood ( Figure 5C and D). However, Rhod-A14 incorporated only into the fibrin fibers of wild-type thrombi and not of factor XIII-deficient thrombi.…”

Tetraspanin Tspan9 regulates platelet collagen receptor GPVI lateral diffusion and activation

“…Tspan9 has relatively strong sequence identity with Tspan4 (55%) and CD53 (43%), for which knockout mice phenotypes have yet to be reported. CD53 is restricted to leukocytes [55] and is absent from platelets [56, 57], so its expression does not appear to overlap with Tspan9. Tspan4 appears to be more widely expressed [58] but not by platelets [56, 57].…”

“…CD53 is restricted to leukocytes [55] and is absent from platelets [56, 57], so its expression does not appear to overlap with Tspan9. Tspan4 appears to be more widely expressed [58] but not by platelets [56, 57]. These expression profiles could explain how a platelet phenotype has been identified in the Tspan9-deficient mice.…”

Modulating platelet reactivity through control of RGS18 availability

“…At least 8 have been detected at the transcript level in platelets, but protein studies suggest that human and mouse platelets express predominantly RGS10 and RGS18. [8][9][10][11] Both proteins are relatively small, consisting primarily of a characteristic RGS domain that interacts with Ga. 12 Each can serve as GTPase-accelerating proteins for Gia and Gqa, but not Gsa. [13][14][15][16][17] RGS18 is primarily expressed in hematopoietic cells 14,16,[18][19][20] whereas RGS10 is widely expressed.…”