There are important questions that remain to be

There are important questions that remain to be answered. How does Rap2a protein affect NF-κB activation? Why do Rap2a silencing and overexpression results to the identical hampered NF-κB activation phenotype? Overexpression experiments have demonstrated that atypical κB-Ras GTPases could inhibit NF-κB activation (Chen et al., 2003; Fenwick et al., 2000; Tago et al., 2010). The major difference between κB-Ras1 and κB-Ras2 and other Ras proteins is the lack of COOH-terminal membrane attachment sequences (CAAX sequences, where “A” is any aliphatic amino Sodium 4-Aminosalicylate and “X” is any amino acid) and the presence of alanine or leucine at position 13 (instead of glycine) and leucine at position 65 (instead of glutamine) (equivalent to positions 12 and 61, respectively, in H-Ras) (Valencia et al., 1991; Macara et al., 1996; Fenwick et al., 2000). These alterations render known Ras proteins into a deregulated, active, GTP-bound conformation and underlies their oncogenicity (Lowy and Willumsen, 1993; Winkler et al., 1997). In the case of Rap2a, though presence of Glycine at position 12, there is a threonine at position 61. Moreover, at the CAAX motif, the second amino acid is non-aliphatic, i.e., charged polar. The observation that κB-Ras1 and 2 harbor V12 and Q61 that render the protein constitutively bound to GTP, and thus maintaining in an activated state, it is possible that GTP-bound Rap2A activated by inflammatory stimuli is a mechanism by which NF-κB activation might be inhibited. Our present study has some limitations. Because of near 90% aminoacid sequence identity among Rap2 members, besides about 60% with Rap1 (see Fig. 8 of Carvalho et al., 2019), one could assume these molecules to serve redundant roles in cells. Experiments with individual silencing or knockout mice may increase our understanding of Rap2 function, as well as the specific functions performed each Rap2a, b or c in macrophages. Additionally, potential off-target effects might complicate the use of siRNA specific for Rap2a. It will be required further studies to assess the expression of Rap2 members and their functions in cells where Rap2a is knocked-down. A number of speculations could arise for the unanticipated signaling role of Rap2a in macrophage activation by TLRs. Rap2 can enhance MAP4K4-induced activation of JNK, and it is suggested as putative effector of Rap2 (Machida et al., 2004). Rap2 activates MAP4K4 and is required for regulation of mechanotransducers target genes CTGF, CYR61, and ANKRD1 (Meng et al., 2018). Moreover, orally delivered siRNA targeting macrophage Map4k4 has been shown to suppress systemic inflammation by regulating the production of inflammatory mediators (Aouadi et al., 2009). One would speculate whether MAP4k4 is a putative effector of Rap2a in mediating cytokines production in TLR-activated macrophages.
Contributions
Declarations of interest
Acknowledgments The authors thank the members of the lab for helpful suggestions and discussion. The authors also thank Josy Hubner for helping with ELISA. BCC is recipient fellow of Coordenação de Aperfeicoamento de Pessoal de Nível Superior (CAPES). AMS is recipient fellow of National Council for Research and Development (CNPq). Funding: This work was supported by grants from brazilian agencies CNPq (National Council for Scientific and Technological Development), CAPES-PNPD (Coordination for higher Education Staff Development, Fundação de Amparo à Pesquisa de Minas Gerais (FAPEMIG), Instituto Nacional de Ciência e Tecnologia de Vacinas (INCTV), and PRPq/UFMG (Pró-reitoria de Pesquisa/ Universidade Federal de Minas Gerais); and by grants to S.G. from the National Institutes of Health (NIH).
The Endolysosomal System of Eukaryotic Cells Eukaryotic cells take advantage of membrane-enclosed compartments to separate enzymatic reactions. Organelles communicate via vesicular carriers that are generated on one organelle, detach, migrate to, and fuse with another organelle. This vesicle-mediated transport is required to deliver specific proteins and lipids to an acceptor organelle 1, 2.