Biology subjects

Burgos, P. V.

Publications and source records attributed to Burgos, P. V..

4 recordsLinked to original sources

bioRxiv2025

Proteasome Inhibition Reduces NBR1 Protein Levels in mATG8-Deficient Cells Independently of Lysosomal Degradation and RAB27A

Autophagy is a lysosome-dependent degradation process that involves autophagosome formation, typically mediated by mammalian ATG8 proteins (mATG8s). Autophagosomes can still form in their absence, suggesting alternative mechanisms. NBR1, a selective autophagy receptor, has been shown to compensate for autophagy defects. Here, we examined NBR1 regulation under proteotoxic stress in mATG8s-deficient HeLa cells. NBR1 levels were elevated in mATG8s knockout (KO) cells under basal conditions but decreased significantly after treatment with the proteasome inhibitor MG132. This reduction was not prevented by lysosomal inhibition with BafA1, indicating a non-lysosomal mechanism. Silencing of RAB27A reduced basal NBR1 levels, and the effects of MG132 were no longer observed, likely due to already diminished NBR1. Remaining NBR1 localized to puncta positive for ubiquitin and the ESCRT-0 component HRS, suggesting involvement of a ubiquitin-dependent endosomal pathway. Overall, our results suggest that under proteotoxic stress and impaired autophagy, cells activae alternative routes, potentially involving unconventional secretion, to regulate NBR1 levels. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC="FIGDIR/small/664987v2_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@10568bcorg.highwire.dtl.DTLVardef@1f3d53org.highwire.dtl.DTLVardef@f33508org.highwire.dtl.DTLVardef@11ec28e_HPS_FORMAT_FIGEXP M_FIG C_FIG Created with BioRender https://BioRender.com/geoh3sz In the absence of mATG8s, NBR1 may be recruited to multivesicular bodies (MVBs) via HRS, for later degradation in lysosomes (left panel). Under proteasomal inhibition (right panel), NBR1 associated with MVBs (with ubiquitinated cargos) is downregulated, independent of lysosomes. Possibly, this downregulation is due to a secretion process, through a RAB27A-independent mechanism.

cell biology↗

bioRxiv2024

The cervical and meningeal lymphatic network as a pathway for retrograde nanoparticle transport to the brain

The meningeal lymphatic vessels have been described as a pathway that transports cerebrospinal fluid and interstitial fluid in a unidirectional manner towards the deep cervical lymph nodes. However, these vessels exhibit anatomical and molecular characteristics typical of initial lymphatic vessels, with the absence of surrounding smooth muscle and few or absent valves. Given its structure, this network could theoretically allow for bidirectional motion. Nevertheless, it has not been assessed as a potential route for nanoparticles to travel from peripheral tissues to the brain. Here we show that extracellular vesicles derived from the B16F10 melanoma cell line, along with superparamagnetic iron oxide nanoparticles, gold nanorods, and Chinese ink nanoparticles can reach the meningeal lymphatic vessels and the brain of C57BL/6 mice after administration within deep cervical lymph nodes in vivo, exclusively through lymphatic structures. Since the functional anatomy of dural lymphatics has been found to be conserved between mice and humans, we expect that our results will encourage further research into the retrograde motion of nanoparticles towards the brain for pharmacological purposes in nanomedicine, as well as to better understand the fluid dynamics in different physiological or neuropathological conditions.

neuroscience↗

bioRxiv2023

Chemo-sEVs release in cisplatin-resistance ovarian cancer cells are regulated by the lysosomal function

Ovarian cancer (OvCa) is an aggressive disease usually treated with cisplatin (CDDP)-based therapy. However, among the different types of cancers treated with CDDP, OvCa commonly develops chemoresistance to this treatment. The small extracellular vesicles (sEVs) play a central role in chemoresistance. In response to chemotherapy, resistant cells secrete sEVs named chemo-sEVs characterized by specific cargo landscape content involved in the transfer of chemoresistance to recipient cells. sEVs encompass a variety of vesicle types, including exosomes, and are formed as intraluminal vesicles (ILVs) within multivesicular endosomes (MVEs). MVEs follow at least two trafficking pathways regulated by RAB GTPase family members; 1) a secretory pathway where MVEs fuse with the plasma membrane (PM) for sEVs secretion, where RAB27A is the most studied; 2) a degradative pathway where MVEs fuse with lysosomes, an event controlled by RAB7. There is growing evidence suggesting that a loss of lysosomal function can increase sEVs secretion; however, whether sEVs secretion and the transfer of CDDP chemoresistance in OvCa is the result of a fine regulation between these two MVEs trafficking pathways is unknown. In this work, we study the status of these two pathways, between CDDP-sensitive (A2780) and CDDP-resistant (A2780cis) OvCa cells. We found A2780cis cells have an increased number of MVEs and ILVs structures, together with higher levels of ESCRTs machinery components and RAB27A, compared to A2780 cells. Moreover, CDDP promotes the secretion of chemo-sEVs in A2780cis cells. Interestingly, chemo-sEVs contain a high number of proteins related to DNA damage response. In addition, we determine A2780cis cells have a poor lysosomal function with reduced levels of RAB7. Surprisingly, silencing of RAB27A in A2780cis cells was found to be sufficient to restore lysosomal function and levels of RAB7 in A2780cis cells, switching into an A2780-like cellular phenotype. Next, we found rapamycin, a potent enhancer of lysosomal function, reduced the secretion of chemo-sEVs. Taken together, these results indicate that the secretion of chemo-sEVs in OvCa cells is determined by the balance between secretory MVEs and MVEs that are destined for lysosomal degradation. Thus, our results suggest that adjusting this balance between these two MVEs trafficking pathways could be a promising strategy for overcoming CDDP chemoresistance in OvCa. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=147 HEIGHT=200 SRC="FIGDIR/small/526974v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@167227corg.highwire.dtl.DTLVardef@91c6forg.highwire.dtl.DTLVardef@29e997org.highwire.dtl.DTLVardef@1a6bebe_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Negative modulation of macroautophagy by HERPUD1 is counteracted by an increased ER-lysosomal network with impact in drug-induced stress cell survival

Macroautophagy and the ubiquitin proteasome system work as an interconnected network in the maintenance of cellular homeostasis. Indeed, efficient activation of macroautophagy upon nutritional deprivation is sustained by degradation of preexisting proteins by the proteasome. However, the specific substrates that are degraded by the proteasome in order to activate macroautophagy are currently unknown. By quantitative proteomic analysis we identified several proteins downregulated in response to starvation but independently of ATG5 expression. Among them, the most significant was HERPUD1, an ER protein of short-half life and a well-known substrate of the proteasome. We found that increased HERPUD1 stability by deletion of its ubiquitin-like domain (UBL) plays a negative role on basal and induced macroautophagy. Moreover, we found it triggers ER expansion by reordering the ER in crystalloid structures, but in the absence of unfolded protein response activation. Surprisingly, we found ER expansion led to an increase in the number and function of lysosomes establishing a tight network with the presence of membrane-contact sites. Importantly, a phosphomimetic S59D mutation within the UBL mimics UBL deletion on its stability and the ER-lysosomal network expansion revealing an increase of cell survival under stress conditions. Altogether, we propose stabilized HERPUD1 downregulates macroautophagy favoring instead a closed interplay between the ER and lysosomes with consequences in drug-cell stress survival. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=198 SRC="FIGDIR/small/447273v2_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1136b35org.highwire.dtl.DTLVardef@1e39b7eorg.highwire.dtl.DTLVardef@1f59211org.highwire.dtl.DTLVardef@148b2b0_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗