Fig 2: (A) Electrophoresis of RT-PCR products for evaluating ERAS mRNA expression in equine tissues. MW: DNA molecular weight marker (100 bp). The PCR was performed both on samples in which reverse transcriptase (RT) was added to the reaction mix and in those without RT (-), using the same amount of RNA. NTC: no template control, omitted RNA template from the reaction mix; C-: PCR negative control. (B) 100% identity between the sequence of the amplicons (Seq PCR cDNA ERas) and the sequences of Equus caballus ES cell expressed Ras (ERAS), mRNA Sequence ID: XM_001493930.3.

Fig 3: (A) Representative WB analysis and (B) ERAS immunoprecipitation (IP) in equine tissues from several animals. WB performed on IP revealed that ERAS interacted with some of proteins involved in mitophagy.

Fig 4: (A) Representative WB analysis and (B) ERAS immunoprecipitation (IP) in equine tissues from several animals. WB analysis carried out on IP revealed that ERAS interacted with some of proteins involved in autophagy.

Fig 5: (A) Western blot (WB) analysis of ERAS protein in equine tissues. C+: WB positive control, ERAS HEK 293T cell transient overexpression lysate. An expected ~25kDa band was shown in all samples. Actin was used as loading control. (B) The values of the densitometric analysis were related to the ERAS protein levels in the positive control (*p < 0.05; **p < 0.01; ***p < 0.001). (C) Alignment between the sequences of the human (NP_853510.1) and equine (XP_001493980.3) ERAS protein showed an 81% identity. The amino acid residues of the epitope against the specific antibody used in the current study have underlined in red. This immunogenic portion of ERAS is composed of 31 amino acid residues (51–81 position) showing a 100% identity between the human and horse epitope.