Biology subjects

Cadena, L. R.

Publications and source records attributed to Cadena, L. R..

6 recordsLinked to original sources

bioRxiv2026

Interconnections with and within the trypanosomal respiratory chain revealed by complexome profiling

Proteins are frequently integrated into multicomponent complexes that execute the elaborate processes supporting life. Thus, a protein's function can only be defined by the company it keeps within a complex. Proteomes provide informative protein inventories but lack information about protein quaternary structures. Complexome profiling (CP) has been transformative in capturing the comprehensive population structure of complexes at a given moment within a cell. We have employed CP to chart the multiprotein complex landscape of two life cycle stages of Trypanosoma brucei. These data have allowed the observation of previously hidden interactions with and within the mitochondrial respiratory chain. We have found (1) an exceptional case of a SLC25 solute transporter that interacts with NADH dehydrogenase, (2) two ATP synthase subunit g paralogs that are intriguingly excluded from the enzyme's dimers, and (3) refined the known composition of ubiquinol:cytochrome c oxidoreductase by addition of missing subunits and removing an incorrectly assigned subunit, which more likely acts to insert the iron-sulfur co-factor into the complex. Further investigation into ubiquinol:cytochrome c oxidoreductase assembly revealed crosstalk between incorporation of its nuclear subunits with mitochondrial translation, possibly facilitating a hitherto unknown quality control mechanism. These discoveries demonstrate the power of our CP data for generation and testing of hypotheses about the mitochondrial and other organellar multiprotein complexes of T. brucei, a protist of medical and evolutionary importance.

cell biology↗

bioRxiv2025

A novel host-encoded protein, ETP2, plays an important role in endosymbiont division in the trypanosomatid Angomonas deanei

A single {beta}-proteobacterial endosymbiont, Candidatus Kinetoplastibacterium crithidii, resides in the cytosol of the trypanosomatid Angomonas deanei, and divides at a defined stage of its hosts cell cycle. This endosymbiont has a highly reduced genome of 0.8 Mb and, notably, has lost most essential genes involved in bacterial cell division, resulting in a loss of division autonomy. It has been previously demonstrated that a host-encoded dynamin-like protein, ETP9, plays an indispensable role in the division of the endosymbiont. In this study, we identified a second nucleus-encoded component of the endosymbiont division machinery, termed ETP2, currently annotated as a hypothetical protein. We observed that ETP2 localizes in a cell cycle-dependent manner at the bacterial division site, alongside the bacterium-encoded FtsZ. Furthermore, we demonstrated that ETP2 plays an important role for timely endosymbiont division and segregation, as a large fraction of cells in an etp2 deletion mutant cell line exhibited either long, filamentous endosymbionts accompanied by severely distorted host cells or host daughter cells lacking endosymbionts. In silico analyses revealed that ETP2 is found exclusively in endosymbiont-harboring trypanosomatids and is most likely an intrinsically disordered protein. Collectively, our data suggests that ETP2, alongside the previously described ETP9, is an integral component of the endosymbiont division machinery. This finding highlights the evolution of a complex host-derived molecular mechanism that exerts tight control over its endosymbiont without requiring gene transfers from the bacterium.

evolutionary biology↗

bioRxiv2024

Characterization of novel and essential kinetoplast-associated proteins in Trypanosoma brucei

The kinetoplast is one of the defining features of kinetoplastid protists and represents a unique concentration of mitochondrial DNA. This subcellular structure is a highly complex assembly of thousands of mutually catenated, circular DNA molecules as well as up to one hundred dedicated proteins. These components work in tandem to replicate and segregate the mitochondrial genome during cellular division, additionally coordinating with the basal body and flagellum through the tripartite attachment complex (TAC) superstructure. Here, we screened the MitoTag localization repository and identified a number of previously undescribed hypothetical proteins exhibiting putative signals within the kinetoplast of Trypanosoma brucei. Through endogenous tagging we verify their association with the kinetoplast or TAC. The essentiality for several of these kinetoplast proteins (KP) was assessed by RNAi knock-downs, revealing that the newly characterized KP56, KP84 and KP86 are indispensable for growth of the procyclic stage. Additionally, KP37, KP56, and KP84 displayed alterations in the abundance of maxicircles or minicircles, while the depletion of KP84 and KP86 resulted in cell cycle alternations. Pulldown assays using the endogenously V5-tagged cell lines identified novel interactors, which were additionally subjected to endogenous tagging for subcellular localization, revealing two additional proteins (KP45 and KP66) with dual localization to the kinetoplast and throughout the mitochondrial lumen. This work represents the most extensive identification of novel KPs to date and provides a methodological pipeline for the characterization of remaining KPs to further understand this intricate molecular structure.

cell biology↗

bioRxiv2024

A novel nabelschnur protein regulates segregation of the kinetoplast DNA in Trypanosoma brucei

The kinetoplast DNA (kDNA), a distinctive arrangement of mitochondrial DNA found in trypanosomatid protists, comprises a concatenated network of minicircles and maxicircles that undergo division and segregation once during each cell cycle. Despite the identification and characterization of numerous proteins involved in kDNA maintenance and replication, its segregation and the formation of the nabelschnur remain poorly understood on a molecular level. This enigmatic filamentous structure, transiently appearing in Trypanosoma brucei, connects the daughter kDNA networks prior to their complete segregation. Here, we characterize TbNAB70, a high mobility group box-like protein localized exclusively to the nabelschnur and the kDNA disc. Our findings demonstrate that TbNAB70 is critical for the segregation, but not replication, of kDNA, a so far unprecedented phenotype. Furthermore, structural predictions suggest that this protein holds the capacity to bind to kDNA illuminating the exact molecular mechanisms of segregation involved. Thus, we propose that TbNAB70 plays a pivotal role in the faithful and efficient segregation of the daughter kDNA networks.

molecular biology↗

bioRxiv2022

The development of intracytoplasmic membranes in alphaproteobacteria involves the conserved mitochondrial crista-developing Mic60 protein

Mitochondrial cristae expand the surface area of respiratory membranes and ultimately allow for the evolutionary scaling of respiration with cell volume across eukaryotes. The discovery of Mic60 homologs among alphaproteobacteria, the closest extant relatives of mitochondria, suggested that cristae might have evolved from bacterial intracytoplasmic membranes (ICMs). Here, we investigated the predicted structure and function of alphaproteobacterial Mic60, and a protein encoded by an adjacent gene Orf52, in two distantly related purple alphaproteobacteria, Rhodobacter sphaeroides and Rhodopseudomonas palustris. In addition, we assessed the potential physical interactors of Mic60 and Orf52 in R. sphaeroides. We show that the three -helices of mitochondrial Mic60s mitofilin domain, as well as its adjacent membrane-binding amphipathic helix, are present in alphaproteobacterial Mic60. The disruption of Mic60 and Orf52 caused photoheterotrophic growth defects, which are most severe under low light conditions, and both their disruption and overexpression led to enlarged ICMs in both studied alphaproteobacteria. We also found that alphaproteobacterial Mic60 physically interacts with BamA, the homolog of Sam50, one of the main physical interactors of eukaryotic Mic60. This interaction, responsible for making contact sites at mitochondrial envelopes, has been conserved in modern alphaproteobacteria despite more than a billion years of evolutionary divergence. Our results suggest a role for Mic60 in photosynthetic ICM development and contact site formation at alphaproteobacterial envelopes. Overall, we provide support for the hypothesis that mitochondrial cristae evolved from alphaproteobacterial ICMs, and therefore have improved our understanding of the nature of the mitochondrial ancestor.

evolutionary biology↗

bioRxiv2021

MICOS and F1FO-ATP synthase crosstalk is a fundamental property of mitochondrial cristae

Mitochondrial cristae are polymorphic invaginations of the inner membrane that are the fabric of cellular respiration. Both the Mitochondrial Contact Site and Cristae Organization System (MICOS) and the FOF1-ATP synthase are vital for sculpting cristae by opposing membrane bending forces. While MICOS promotes negative curvature at cristae junctions, dimeric FOF1- ATP synthase is crucial for positive curvature at cristae rims. Crosstalk between these two complexes has been observed in bakers yeast, the model organism of the Opisthokonta supergroup. Here, we report that this property is conserved in Trypanosoma brucei, a member of the Discoba supergroup that separated from Opisthokonta [~]2 billion years ago. Specifically, one of the paralogs of the core MICOS subunit Mic10 interacts with dimeric F1FO-ATP synthase, whereas the other core Mic60 subunit has a counteractive effect on F1FO- ATP synthase oligomerization. This is evocative of the nature of MICOS-F1FO-ATP synthase crosstalk in yeast, which is remarkable given the diversification these two complexes have undergone during almost 2 eons of independent evolution. Furthermore, we identified a highly diverged trypanosome homolog of subunit e, which is essential for the stability of F1FO-ATP synthase dimers in yeast. Just like subunit e, it is preferentially associated with dimers, interacts with Mic10 and its silencing results in severe defects to cristae and disintegration of F1FO-ATP synthase dimers. Our findings indicate that crosstalk between MICOS and dimeric F1FO-ATP synthase is a fundamental property impacting cristae shape throughout eukaryotes. ImportanceMitochondria have undergone profound diversification in separate lineages that have radiated since the last common ancestor of eukaryotes some eons ago. Most eukaryotes are unicellular protists, including etiological agents of infectious diseases like Trypanosoma brucei. Thus, the study of a broad range of protists can reveal fundamental features shared by all eukaryotes and lineage-specific innovations. Here we report that two different protein complexes, MICOS and F1FO-ATP synthase, known to affect mitochondrial architecture, undergo crosstalk in T. brucei, just as in bakers yeast. This is remarkable considering that these complexes have otherwise undergone many changes during their almost two billion years of independent evolution. Thus, this crosstalk is a fundamental property needed to maintain proper mitochondrial structure even if the constituent players considerably diverged.

cell biology↗