Revisiting the intraperoxisomal pathway of mammalian PEX7

Newly synthesized peroxisomal proteins containing a cleavable type 2 targeting signal (PTS2) are transported to the peroxisome by a cytosolic PEX5-PEX7 complex. There, the trimeric complex becomes inserted into the peroxisomal membrane docking/translocation machinery (DTM), a step that leads to the translocation of the cargo into the organelle matrix. Previous work suggests that PEX5 is retained at the DTM during all the steps occurring at the peroxisome but whether the same applies to PEX7 was unknown. By subjecting different pre-assembled trimeric PEX5-PEX7-PTS2 complexes to in vitro co-import/export assays we found that the export competence of peroxisomal PEX7 is largely determined by the PEX5 molecule that transported it to the peroxisome. This finding suggests that PEX7 is also retained at the DTM during the peroxisomal steps and implies that cargo proteins are released into the organelle matrix by DTM-embedded PEX7. The release step does not depend on PTS2 cleavage. Rather, our data suggest that insertion of the trimeric PEX5-PEX7-PTS2 protein complex into the DTM is probably accompanied by conformational alterations in PEX5 to allow release of the PTS2 protein into the organelle matrix.This work was funded by FEDER funds through the Operational Competitiveness Programme, COMPETE,and by National Funds through FCT, Fundacao para a Ciencia e a Tecnologia, under the project FCOMP-01-0124-FEDER-022718 (Pest-C/SAU/LA0002/2011) and FCOMP-01-0124-FEDER-019731 (PTDC/BIABCM/118577/2010). T.A.R. and C.P.G were supported by Fundacao para a Ciencia e a Tecnologia,Programa Operacional Potencial Humano do QREN and Fundo Social Europeu

A mechanistic perspective on pex1 and pex6, two aaa+ proteins of the peroxisomal protein import machinery

In contrast to many protein translocases that use ATP or GTP hydrolysis as the driving force to transport proteins across biological membranes, the peroxisomal matrix protein import machinery relies on a regulated self-assembly mechanism for this purpose and uses ATP hydrolysis only to reset its components. The ATP-dependent protein complex in charge of resetting this machinery—the Receptor Export Module (REM)—comprises two members of the “ATPases Associated with diverse cellular Activities” (AAA+) family, PEX1 and PEX6, and a membrane protein that anchors the ATPases to the organelle membrane. In recent years, a large amount of data on the structure/function of the REM complex has become available. Here, we discuss the main findings and their mechanistic implications.This work was financed by FEDER—Fundo Europeu de Desenvolvimento Regional funds through the COMPETE 2020—Operacional Programme for Competitiveness and Internationalisation (POCI), Portugal 2020, and by Portuguese funds through FCT—Fundação para a Ciência e a Tecnologia/Ministério da Ciência, Tecnologia e Ensino Superior in the framework of the project PTDC/BEX-BCM/2311/2014 (POCI-01-0145-FEDER-016613) and the project “Institute for Research and Innovation in Health Sciences” (POCI-01-0145-FEDER-007274). This work is a result of the project NORTE-01-0145-FEDER-000008—Porto Neurosciences and Neurologic Disease Research Initiative at I3S, supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (FEDER). A.B.-B., A.G.P., M.J.F., T.F. and T.A.R. are supported by Fundação para a Ciência e Tecnologia, Programa Operacional Potencial Humano do QREN, and Fundo Social Europeu

The N terminus of the peroxisomal cycling receptor, Pex5p, is required for redirecting the peroxisome-associated peroxin back to the cytosol

Most newly synthesized peroxisomal matrix proteins are transported to the organelle by Pex5p, a remarkable multidomain protein involved in an intricate network of transient protein-protein interactions. Presently, our knowledge regarding the structure/function of amino acid residues 118 to the very last residue of mammalian Pex5p is quite vast. Indeed, the cargo-protein receptor domain as well as the binding sites for several peroxins have all been mapped to this region of Pex5p. In contrast, structural/functional data regarding the first 117 amino acid residues of Pex5p are still scarce. Here we show that a truncated Pex5p lacking the first 110 amino acid residues (DeltaN110-Pex5p) displays exactly the peroxisomal import properties of the full-length peroxin implying that this N-terminal domain is involved neither in cargo-protein binding nor in the docking/translocation step of the Pex5p-cargo protein complex at the peroxisomal membrane. However, the ATP-dependent export step of DeltaN110-Pex5p from the peroxisomal membrane is completely blocked, a phenomenon that was also observed for a Pex5p version lacking just the first 17 amino acid residues but not for a truncated protein comprising amino acid residues 1-324 of Pex5p. By exploring the unique properties of DeltaN110-Pex5p, the effect of temperature on the import/export kinetics of Pex5p was characterized. Our data indicate that the export step of Pex5p from the peroxisomal compartment ( in contrast with its insertion into the organelle membrane) is highly dependent on the temperature

Primary biliary cirrhosis in a rheumatoid arthritis patient treated with rituximab, a case-based review

Primary biliary cirrhosis (PBC) is an autoimmune disease in which intrahepatic bile ducts are targeted by an immune-mediated injury. This disease tends to progress to fibrosis and cirrhosis with hepatic failure. The authors report a case of a 50-year-old rheumatoid arthritis (RA) patient, with erosions and seropositive for rheumatoid factor and anti-citrullinated peptide antibodies, with 18 years disease duration refractory to prednisolone and several disease-modifying antirheumatic drugs, either conventional or biological (adalimumab and etanercept). In April 2007, she started therapy with rituximab (RTX) with good European League Against Rheumatism response achieved 9 months later. In June 2008, she was admitted with intrahepatic cholestasis, steatorrhea, and spontaneous fractures of various ribs. After excluding cholelitiasis, as well as infectious and neoplastic diseases a liver biopsy was performed that was compatible with the diagnosis of PBC. The antinuclear antibodies (1/160) were positive as well as the antimitochondrial antibodies (1/640). Other antibodies were negative such as anti-SSA and anti-SSB. Afterwards, the patient started ursodesoxycholic acid 15 mg kg(-1) day(-1) with progressive improvement of cholestatic markers. A labial salivary gland biopsy was performed and showed findings compatible with the concomitant diagnosis of Sjögren's syndrome. Based on this clinical report, a detailed review of the clinical aspects of PBC is presented as well as its association with other immune-mediated inflammatory diseases, particularly, with RA

Bone Histomorphometry Revisited

Bone histomorphometry is defined as a quantitative evaluation of bone micro architecture, remodelling and metabolism. Bone metabolic assessment is based on a dynamic process, which provides data on bone matrix formation rate by incorporating a tetracycline compound. In the static evaluation, samples are stained and a semi-automatic technique is applied in order to obtain bone microarchitectural parameters such as trabecular area, perimeter and width. These parameters are in 2D, but they can be extrapolated into 3D, applying a stereological formula. Histomorphometry can be applied to different areas; however, in recent decades it has been a relevant tool in monitoring the effect of drug administration in bone. The main challenge for the future will be the development of noninvasive methods that can give similar information. In the herein review paper we will discuss the general principles and main applications of bone histomorphometry

Impact of a legumes diet on the human gut microbiome articulated with fecal and plasma metabolomes: A pilot study

Background & aims: Legumes intake is known to be associated with several health benefits the origins of which is still a matter of debate. This paper addresses a pilot small cohort to probe for metabolic aspects of the interplay between legumes intake, human metabolism and gut microbiota. Methods: Untargeted nuclear magnetic resonance (NMR) metabolomics of blood plasma and fecal extracts was carried out, in tandem with qPCR analysis of feces, to assess the impact of an 8-week pilot legumes diet intervention on the fecal and plasma metabolomes and gut microbiota of 19 subjects. Results: While the high inter-individual variability hindered the detection of statistically significant changes in the gut microbiome, increased fecal glucose and decreased threonine levels were noted. Correlation analysis between the microbiome and fecal metabolome lead to putative hypotheses regarding the metabolic activities of prevalent bacteria groups (Clostridium leptum subgroup, Roseburia spp., and Faecalibacterium prausnitzii). These included elevated fecal glucose as a preferential energy source, the involvement of valerate/isovalerate and reduced protein degradation in gut microbiota. Plasma metabolomics advanced mannose and betaine as potential markers of legume intake and unveiled a decrease in formate and ketone bodies, the latter suggesting improved energy utilization through legume carbohydrates. Amino acid metabolism was also apparently affected, as suggested by lowered urea, histidine and threonine levels. Conclusions: Despite the high inter-individual gut microbiome variability characterizing the small cohort addressed, combination of microbiological measurements and untargeted metabolomics unveiled several metabolic effects putatively related to legumes intake. If confirmed in larger cohorts, our findings will support the inclusion of legumes in diets and contribute valuable new insight into the origins of associated health benefits. © 2024 The Author(s)The authors acknowledge the help of Joana Ferrão Silveira, MSc and Kimhoung Say, MSc during both the food intervention and data collection procedures, as well as the food company EUREST for providing the trial meals. This work was supported by National Funds from FCT - Fundação para a Ciência e a Tecnologia through project UIDB/50016/2020 and by the project “Transition paths to sustainable legume-based systems in Europe” (TRUE) which has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No. 727973. HF would like to acknowledge FCT for doctoral grant ref. SFRH/BDE/132240/2017. AMG acknowledges the project CICECO-Aveiro Institute of Materials, UIDB/50011/2020 (DOI 10.54499/UIDB/50011/2020), UIDP/50011/2020 (DOI 10.54499/UIDP/50011/2020) & LA/P/0006/2020 (DOI 10.54499/LA/P/0006/2020), financed by national funds through the FCT/MCTES (PIDDAC). The NMR spectrometer is part of the National NMR Network (PTNMR) and are partially supported by Infrastructure Project Nº 022161 (co-financed by FEDER through COMPETE 2020, POCI and PORL and FCT through PIDDAC)

The first minutes in the life of a peroxisomal matrix protein

In the field of intracellular protein sorting, peroxisomes are most famous by their capacity to import oligomeric proteins. The data supporting this remarkable property are abundant and, understandably, have inspired a variety of hypothetical models on how newly synthesized (cytosolic) proteins reach the peroxisome matrix. However, there is also accumulating evidence suggesting that many peroxisomal oligomeric proteins actually arrive at the peroxisome still as monomers. In support of this idea, recent data suggest that PEX5, the shuttling receptor for peroxisomal matrix proteins, is also a chaperone/holdase, binding newly synthesized peroxisomal proteins in the cytosol and blocking their oligomerization. Here we review the data behind these two different perspectives and discuss their mechanistic implications on this protein sorting pathway. This article is part of a Special Issue entitled: Peroxisomes edited by Ralf Erdmann.This work was supported by national funds through FCT – Fundação para a Ciência e a Tecnologia/MEC-Ministério da Educação e Ciência and when applicable co-funded by FEDER funds within the partnership agreement PT2020 related with the research unit number 4293; and by the project FCOMP-01-0124-FEDER-019731 (PTDC/BIABCM/118577/2010) funded by national funds through FCT and co-funded by Fundo Europeu de Desenvolvimento Regional (FEDER) through the Operation- alCompetitiveness Programme(COMPETE).A. F.D., T.F., T.A.R. and C. P. G. were supported by FCT, Programa Operacional Potencial Humano (POPH) do Quadro de Referência Estratégico Nacional (QREN), and Fundo Social Europeu (FSE)

BioRePortAP, an electronic clinical record coupled with a database: an example of its use in a single centre

AIMS: To evaluate the efficacy and safety of the treatment of psoriatic arthritis (PsA) patients with tumor necrosis factor (TNF) antagonists in the Rheumatology Department of Hospital de Santa Maria using the BioRePortAP. METHODS: The Portuguese Society of Rheumatology (SPR) developed an electronic medical chart coupled with a database for the follow up of PsA patients, the BioRePortAP, which was launched in May 2009. This evaluation was based on all the PsA patients that were on active treatment with TNF antagonists in September 2009 and were registered in the BioRePortAP. All the previous data on these patients were introduced in BioRePortAP using the prospective paper based follow up protocol that this Department was using since 1999. Only patients with more than 9 months of treatment were analyzed. RESULTS: Forty-two patients with PsA, actively treated with anti-TNF agents in September 2009, for at least 9 months, were analyzed in BioRePortAP. Twenty-three patients were male (55%) and nineteen were female (45%). The average age of these patients was 49.8+/-10.9 years old, the average disease duration was of 10.7+/-5.6 years and the mean duration of biological therapy was of 37.8+/-27.8 months. For the 81% of patients with peripheral joint disease there was a mean reduction of more than 80% in the swollen and tender joint counts, and almost 50% in the health assessment questionnaire (HAQ) value. In the 19% of the patients with axial involvement the reduction of BASDAI and BASFI was not statistically significative. On top of that, PASI score suffered a reduction of 64%. Fourteen patients (33.3%) had to switch their TNF antagonist treatment. 58.8% of the switches were due to adverse effects and 41.2% due to therapy failure. Regarding the 56 adverse reactions registered, only one was a severe reaction. The remaining adverse reactions were not severe and 67% of them were due to infections. DISCUSSION: The results of this first report of the use of the BioRePortAP in clinical practice confirm the efficacy and safety of TNF antagonist treatment in PsA. The results shown here elucidate the potential applications of BioRePortAP as a tool for efficacy and safety assessment of PsA patients treated with biotechnological drugs