Search

Muscarinic acetylcholine receptors are prototypical G protein-coupled receptors (GPCRs), members of a large family of seven transmembrane receptors that mediates a wide variety of extracellular signals. We show here, for the first time in a GPCR, that the carboxyl terminal fragment of the muscarinic M2receptor, containing the transmembrane regions 6 and 7 (M2tail), is expressed by virtue of an internal ribosome entry site located in the third intracellular loop. Single cell imaging reveals that M2tail, whose expression is upregulated in cells undergoing integrated stress response, does not follow the normal route to the plasma membrane, but is almost exclusively sorted to the mitochondria inner membrane: here it controls oxygen consumption, cell proliferation and the formation of reactive oxygen species by reducing oxidative phosphorylation. Crispr/Cas9 editing of the key methionine where cap-independent translation begins in human induced pluripotent stem cells, reveals the physiological role of this process in influencing cell proliferation and oxygen consumption at the endogenous level. The expression of the C-terminal domain of a GPCR, capable of regulating mitochondrial function, constitutes a hitherto unknown mechanism notably unrelated to its canonical signalling function as a GPCR at the plasma membrane. This work thus highlights a potential novel mechanism thar cells may use for controlling their metabolism under variable environmental conditions, notably as a negative regulator of cell respiration.

Spatially controlled, cargo-specific endocytosis is essential for development, tissue homeostasis and cancer invasion. Unlike cargo-specific clathrin-mediated endocytosis, the clathrin- and dynamin-independent endocytic pathway (CLIC-GEEC, CG pathway) is considered a bulk internalization route for the fluid phase, glycosylated membrane proteins and lipids. While the core molecular players of CG-endocytosis have been recently defined, evidence of cargo-specific adaptors or selective uptake of proteins for the pathway are lacking. Here we identify the actin-binding protein Swiprosin-1 (Swip1, EFHD2) as a cargo-specific adaptor for CG-endocytosis. Swip1 couples active Rab21-associated integrins with key components of the CG-endocytic machinery-Arf1, IRSp53 and actin-and is critical for integrin endocytosis. Through this function, Swip1 supports integrin-dependent cancer-cell migration and invasion, and is a negative prognostic marker in breast cancer. Our results demonstrate a previously unknown cargo selectivity for the CG pathway and a role for specific adaptors in recruitment into this endocytic route.

How do cells maintain relative proportions of protein complex components? Advances in quantitative, genome-wide measurements have begun to shed light onto the roles of protein synthesis and degradation in establishing the precise proportions in living cells: on the one hand, ribosome profiling studies indicate that proteins are already produced in the correct relative proportions. On the other hand, proteomic studies found that many complexes contain subunits that are made in excess and subsequently degraded. Here, we discuss these seemingly contradictory findings, emerging principles, and remaining open questions. We conclude that establishing precise protein levels involves both coordinated synthesis and post-translational fine-tuning via protein degradation.

Calcium influx through the voltage-gated L-type calcium channels (LTCC) mediates a wide range of physiological processes from contraction to secretion. Despite extensive research on regulation of LTCC conductance by PKA phosphorylation in response to β-adrenergic stimulation, the science remains incomplete. Here, we show that Wnt11, a non-canonical Wnt ligand, through its G protein-coupled receptor (GPCR) Fzd7 attenuates the LTCC conductance by preventing the proteolytic processing of its C terminus. This is mediated across species by protein kinase A (PKA), which is compartmentalized by A-kinase anchoring proteins (AKAP). Systematic analysis of all AKAP family members revealed AKAP2 anchoring of PKA is central to the Wnt11-dependent regulation of the channel. The identified Wnt11/AKAP2/PKA signalosome is required for heart development, controlling the intercellular electrical coupling in the developing zebrafish heart. Altogether, our data revealed Wnt11/Fzd7 signaling via AKAP2/PKA as a conserved alternative GPCR system regulating Ca2+homeostasis.

The principle of shotgun proteomics is to use peptide mass spectra in order to identify corresponding sequences in a protein database. The quality of peptide and protein identification and quantification critically depends on the sensitivity and specificity of this assignment process. Many peptides in proteomic samples carry biochemical modifications, and a large fraction of unassigned spectra arise from modified peptides. Spectra derived from modified peptides can erroneously be assigned to wrong amino acid sequences. However, the impact of this problem on proteomic data has not yet been investigated systematically. Here we use combinations of different database searches to show that modified peptides can be responsible for 20–50% of false positive identifications in deep proteomic data sets. These false positive hits are particularly problematic as they have significantly higher scores and higher intensities than other false positive matches. Furthermore, these wrong peptide assignments lead to hundreds of false protein identifications and systematic biases in protein quantification. We devise a “cleaned search” strategy to address this problem and show that this considerably improves the sensitivity and specificity of proteomic data. In summary, we show that modified peptides cause systematic errors in peptide and protein identification and quantification and should therefore be considered to further improve the quality of proteomic data annotation.

Targeted proteomic approaches like selected reaction monitoring (SRM) and parallel reaction monitoring (PRM) are increasingly popular because they enable sensitive and rapid analysis of a preselected set of proteins. However, developing targeted assays is tedious and requires the selection, synthesis and mass spectrometric analysis of candidate peptides before the actual samples can be analyzed. The SRMatlas and ProteomeTools projects recently published fragmentation spectra of synthetic peptides covering the entire human proteome. These datasets provide very valuable resources. However, extracting the relevant data for selected proteins of interest is not straightforward. For example, developing scheduled acquisition methods (i.e. analyzing specific peptides in defined elution time windows) is complicated and requires adjustments to specific chromatographic conditions employed. Here, we present Picky: a fast and easy to use online tool to design scheduled PRM/SRM assays (https://picky.mdc-berlin.de). Picky (i) automatically generates optimized and scheduled SRM/PRM assays for proteins of interest and (ii) provides means to validate the data via known fragmentation spectra of corresponding synthetic peptides.

Many disease-causing missense mutations affect intrinsically disordered regions (IDRs) of proteins, but the molecular mechanism of their pathogenicity is enigmatic. Here, we employ a peptide-based proteomic screen to investigate the impact of mutations in IDRs on protein-protein interactions. We find that mutations in disordered cytosolic regions of three transmembrane proteins (GLUT1, ITPR1, and CACNA1H) lead to an increased clathrin binding. All three mutations create dileucine motifs known to mediate clathrin-dependent trafficking. Follow-up experiments on GLUT1 (SLC2A1), the glucose transporter causative of GLUT1 deficiency syndrome, revealed that the mutated protein mislocalizes to intracellular compartments. Mutant GLUT1 interacts with adaptor proteins (APs) in vitro, and knocking down AP-2 reverts the cellular mislocalization and restores glucose transport. A systematic analysis of other known disease-causing variants revealed a significant and specific overrepresentation of gained dileucine motifs in structurally disordered cytosolic domains of transmembrane proteins. Thus, several mutations in disordered regions appear to cause "dileucineopathies."

Emerging evidence indicates that heterogeneity in ribosome composition can give rise to specialized functions. Until now, research mainly focused on differences in core ribosomal proteins and associated factors. The effect of posttranslational modifications has not been studied systematically. Analyzing ribosome heterogeneity is challenging because individual proteins can be part of different subcomplexes (40S, 60S, 80S, and polysomes). Here we develop polysome proteome profiling to obtain unbiased proteomic maps across ribosomal subcomplexes. Our method combines extensive fractionation by sucrose gradient centrifugation with quantitative mass spectrometry. The high resolution of the profiles allows us to assign proteins to specific subcomplexes. Phosphoproteomics on the fractions reveals that phosphorylation of serine 38 in RPL12/uL11, a known mitotic CDK1 substrate, is strongly depleted in polysomes. Follow-up experiments confirm that RPL12/uL11 phosphorylation regulates the translation of specific subsets of mRNAs during mitosis. Together, our results show that posttranslational modification of ribosomal proteins can regulate translation.

Targeted proteomic approaches like selected reaction monitoring (SRM) and parallel reaction monitoring (PRM) are increasingly popular because they enable sensitive and rapid analysis of a preselected set of proteins1-3. However, developing targeted assays is tedious and requires the selection, synthesis and mass spectrometric analysis of candidate peptides before the actual samples can be analyzed. The SRMatlas and ProteomeTools projects recently published fragmentation spectra of synthetic peptides covering the entire human proteome4,5. These datasets provide very valuable resources. However, extracting the relevant data for selected proteins of interest is not straightforward. For example, developing scheduled acquisition methods (i.e. analyzing specific peptides in defined elution time windows) is complicated and requires adjustments to specific chromatographic conditions employed. Moreover, the number of peptide candidates to be targeted in parallel often exceeds the analytical abilities of the mass spectrometer. In this case, the key question is which peptides can be omitted without losing too much information. Ideally, a method design tool would automatically select the most informative peptides in each retention time window. Until now, none of the available tools automatically generates such optimized scheduled SRM and PRM methods (Figure S1).

Clathrin-mediated endocytosis is the major mechanism for eukaryotic plasma membrane-based proteome turn-over. In plants, clathrin-mediated endocytosis is essential for physiology and development, but the identification and organization of the machinery operating this process remains largely obscure. Here, we identified an eight-core-component protein complex, the TPLATE complex, essential for plant growth via its role as major adaptor module for clathrin-mediated endocytosis. This complex consists of evolutionarily unique proteins that associate closely with core endocytic elements. The TPLATE complex is recruited as dynamic foci at the plasma membrane preceding recruitment of adaptor protein complex 2, clathrin, and dynamin-related proteins. Reduced function of different complex components severely impaired internalization of assorted endocytic cargoes, demonstrating its pivotal role in clathrin-mediated endocytosis. Taken together, the TPLATE complex is an early endocytic module representing a unique evolutionary plant adaptation of the canonical eukaryotic pathway for clathrin-mediated endocytosis.

Although Rho GTPases are essential molecular switches involved in many cellular processes, an unbiased experimental comparison of their interaction partners was not yet performed. Here, we develop quantitative GTPase affinity purification (qGAP) to systematically identify interaction partners of six Rho GTPases (Cdc42, Rac1, RhoA, RhoB, RhoC, and RhoD), depending on their nucleotide loading state. The method works with cell line or tissue-derived protein lysates in combination with SILAC-based or label-free quantification, respectively. We demonstrate that qGAP identifies known and novel binding partners that can be validated in an independent assay. Our interaction network for six Rho GTPases contains many novel binding partners, reveals highly promiscuous interaction of several effectors, and mirrors evolutionary relationships among Rho GTPases.

The large-scale analysis of thousands of proteins under various experimental conditions or in mutant lines has gained more and more importance in hypothesis-driven scientific research and systems biology in the past years. Quantitative analysis by large scale proteomics using modern mass spectrometry usually results in long lists of peptide ion intensities. The main interest for most researchers, however, is to draw conclusions on the protein level. Postprocessing and combining peptide intensities of a proteomic data set requires expert knowledge, and the often repetitive and standardized manual calculations can be time-consuming. The analysis of complex samples can result in very large data sets (lists with several 1000s to 100,000 entries of different peptides) that cannot easily be analyzed using standard spreadsheet programs. To improve speed and consistency of the data analysis of LC-MS derived proteomic data, we developed cRacker. cRacker is an R-based program for automated downstream proteomic data analysis including data normalization strategies for metabolic labeling and label free quantitation. In addition, cRacker includes basic statistical analysis, such as clustering of data, or ANOVA and t tests for comparison between treatments. Results are presented in editable graphic formats and in list files.

Plants have an unusual plastid-localized NADP-thioredoxin reductase C (NTRC) containing both an NADP-thioredoxin reductase (NTR) and a thioredoxin (Trx) domain in a single polypeptide. Although NTRC is known to supply reductant for detoxifying hydrogen peroxide in the dark, its other functions are unknown. We now report that NTRC plays a previously unrecognized role in the redox regulation of ADP-glucose pyrophosphorylase (AGPase), a central enzyme of starch synthesis. When supplied NADPH, NTRC activated AGPase in vitro in a redox reaction that required the active site cysteines of both domains of the enzyme. In leaves, AGPase was activated in planta either by light or external feeding of sucrose in the dark. Leaves of an Arabidopsis NTRC KO mutant showed a decrease both in the extent of redox activation of AGPase and in the enhancement of starch synthesis either in the light (by 40-60%) or in the dark after treatment with external sucrose (by almost 100%). The light-dependent activation of AGPase in isolated chloroplasts, by contrast, was unaffected. In nonphotosynthetic tissue (roots), KO of NTRC decreased redox activation of AGPase and starch synthesis in response to light or external sucrose by almost 90%. The results provide biochemical and genetic evidence for a role of NTRC in regulating starch synthesis in response to either light or sucrose. The data also suggest that the Trx domain of NTRC and, to a lesser extent, free Trxs linked to ferredoxin enable amyloplasts of distant sink tissues to sense light used in photosynthesis by leaf chloroplasts and adjust heterotrophic starch synthesis accordingly. Ministerio de Educación y Ciencia BIO2007– 60644 Junta de Andalucía P06-CVI-01578 y CVI-182

RNA sequencing protocols allow for quantifying gene expression regulation at each individual step, from transcription to protein synthesis. Ribosome Profiling (Ribo-seq) maps the positions of translating ribosomes over the entire transcriptome. Despite its great potential, a rigorous statistical approach to identify translated regions by means of the characteristic three-nucleotide periodicity of Ribo-seq data is not yet available. To fill this gap, we developed RiboTaper, which quantifies the significance of periodic Ribo-seq reads via spectral analysis methods. We applied RiboTaper on newly generated, deep Ribo-seq data in HEK293 cells, to derive an extensive map of translation that covers Open Reading Frame (ORF) annotations for more than 11,000 protein- coding genes. We also find distinct ribosomal signatures for several hundred detected upstream ORFs and ORFs in annotated non-coding genes (ncORFs). Mass spectrometry data confirms that RiboTaper achieves excellent coverage of the cellular proteome and validates dozens of novel peptide products. Collectively, RiboTaper (available at https://ohlerlab.mdc-berlin.de/software/ ) is a powerful method for comprehensive de novo identification of actively used ORFs in the human genome.

The plasma membrane is an important compartment that undergoes dynamic changes in composition upon external or internal stimuli. The dynamic subcompartmentation of proteins in ordered low-density (DRM) and disordered high-density (DSM) membrane phases is hypothesized to require interactions with cytoskeletal components. Here, we systematically analyzed the effects of actin or tubulin disruption on the distribution of proteins between membrane density phases. We used a proteomic screen to identify candidate proteins with altered submembrane location, followed by biochemical or cell biological characterization in Arabidopsis thaliana. We found that several proteins, such as plasma membrane ATPases, receptor kinases, or remorins resulted in a differential distribution between membrane density phases upon cytoskeletal disruption. Moreover, in most cases, contrasting effects were observed: Disruption of actin filaments largely led to a redistribution of proteins from DRM to DSM membrane fractions while disruption of tubulins resulted in general depletion of proteins from the membranes. We conclude that actin filaments are necessary for dynamic movement of proteins between different membrane phases and that microtubules are not necessarily important for formation of microdomains as such, but rather they may control the protein amount present in the membrane phases.

There is increasing evidence that non-annotated short open reading frames (sORFs) can encode functional micropeptides, but computational identification remains challenging. We expand our published method and predict conserved sORFs in human, mouse, zebrafish, fruit fly and the nematode C. elegans. Isolating specific conservation signatures indicative of purifying selection on encoded amino acid sequence, we identify about 2000 novel sORFs in the untranslated regions of canonical mRNAs or on transcripts annotated as non-coding. Predicted sORFs show stronger conservation signatures than those identified in previous studies and are sometimes conserved over large evolutionary distances. Encoded peptides have little homology to known proteins and are enriched in disordered regions and short interaction motifs. Published ribosome profiling data indicate translation for more than 100 of novel sORFs, and mass spectrometry data gives peptidomic evidence for more than 70 novel candidates. We thus provide a catalog of conserved micropeptides for functional validation in vivo.

The plasma membrane is an important organelle providing structure, signaling and transport as major biological functions. Being composed of lipids and proteins with different physicochemical properties, the biological functions of membranes depend on specific protein-protein and protein-lipid interactions. Interactions of proteins with their specific sterol and lipid environment were shown to be important factors for protein recruitment into sub-compartmental structures of the plasma membrane. System-wide implications of altered endogenous sterol levels for membrane functions in living cells were not studied in higher plant cells. In particular, little is known how alterations in membrane sterol composition affect protein and lipid organization and interaction within membranes. Here, we conducted a comparative analysis of the plasma membrane protein and lipid composition in Arabidopsis sterol-biosynthesis mutants smt1 and ugt80A2;B1. smt1 shows general alterations in sterol composition while ugt80A2;B1 is significantly impaired in sterol glycosylation. By systematically analyzing different cellular fractions and combining proteomic with lipidomic data we were able to reveal contrasting alterations in lipid-protein interactions in both mutants, with resulting differential changes in plasma membrane signaling status.

Chelidonium majus and Corydalis cava are phylogenetically closely related (Papaveraceae family). The medicinal and pharmaceutical interest in these plants is based on their synthesis of pharmaceutically important compounds, such as alkaloids, flavonoids, phenolic acids and proteins. C. majus shoot and C. cava tuber extracts have been used in traditional folk medicine to treat many diseases, such as fungal, bacterial and viral infections, liver disorders, fever, post-traumatic, colic, abdominal and menstrual pains and even cancer. This study attempts to perform a global comparative proteomic analysis of pharmacologically important extracts from these two closely related unsequenced plant species to gain insights into the protein basis of these plant organs and to compare their common and specific proteomic compositions. We used a shotgun proteomic approach combined with label-free protein quantitation according to the exponentially modified protein abundance index (emPAI). In total, a mean number of 228 protein identification results were recorded in C. cava tuber extracts and about 1240 in C. majus shoot extracts. Comparative analysis revealed a similar stress and defense-related protein composition of pharmacologically active plant species and showed the presence of different pathogenesis-related and low molecular inducible antimicrobial peptides. These findings could form the basis for further elucidation of the mechanism of the strong pharmacological activities of these medicinal plant extracts.

During the last decade, research on plasma membrane focused increasingly on the analysis of so-called microdomains. It has been shown that function of many membrane-associated proteins involved in signaling and transport depends on their conditional segregation within sterol-enriched membrane domains. High throughput proteomic analysis of sterol-protein interactions are often based on analyzing detergent resistant membrane fraction enriched in sterols and associated proteins, which also contain proteins from these microdomain structures. Most studies so far focused exclusively on the characterization of detergent resistant membrane protein composition and abundances. This approach has received some criticism because of its unspecificity and many co-purifying proteins. In this study, by a label-free quantitation approach, we extended the characterization of membrane microdomains by particularly studying distributions of each protein between detergent resistant membrane and detergent-soluble fractions (DSF). This approach allows a more stringent definition of dynamic processes between different membrane phases and provides a means of identification of co-purifying proteins. We developed a random sampling algorithm, called Unicorn, allowing for robust statistical testing of alterations in the protein distribution ratios of the two different fractions. Unicorn was validated on proteomic data from methyl-β-cyclodextrin treated plasma membranes and the sterol biosynthesis mutant smt1. Both, chemical treatment and sterol-biosynthesis mutation affected similar protein classes in their membrane phase distribution and particularly proteins with signaling and transport functions.

Quantitative comparative analyses of protein abundances using peptide ion intensities and their modifications have become a widely used technique in studying various biological questions. In the past years, several methods for quantitative proteomics were established using stable-isotope labeling and label-free approaches. We systematically evaluated the application of reference protein normalization (RPN) for proteomic experiments using a high mass accuracy LC-MS/MS platform. In RPN all sample peptide intensities were normalized to an average protein intensity of a spiked reference protein. The main advantage of this method is that it avoids fraction of total based relative analysis of proteomic data, which is often very much dependent on sample complexity. We could show that reference protein ion intensity sums are sufficiently reproducible to ensure a reliable normalization. We validated the RPN strategy by analyzing changes in protein abundances induced by nutrient starvation in Arabidopsis thaliana. Beyond that, we provide a principle guideline for determining optimal combination of sample protein and reference protein load on individual LC-MS/MS systems.

As noninvasively accessible body fluid, saliva is of growing interest in diagnostics. To exemplify the diagnostic potential of saliva, we used a mass spectrometry-based approach to gain insights into adaptive physiological processes underlying long-lasting endurance work load in a case study. Saliva was collected from male and female athlete at four diurnal time points throughout a 1060 km nonstop cycling event. Total sampling time covered 180 h comprising 62 h of endurance cycling as well as reference samples taken over 3 days before the event, and over 2 days after. Altogether, 1405 proteins and 62 metabolites were identified in these saliva samples, of which 203 could be quantified across the majority of the sampling time points. Many proteins show clear diurnal abundance patterns in saliva. In many cases, these patterns were disturbed and altered by the long-term endurance stress. During the stress phase, metabolites of energy mobilization, such as creatinine and glucose were of high abundance, as well as metabolites with antioxidant functions. Lysozyme, amylase, and proteins with redox-regulatory function showed significant increase in average abundance during work phase compared to rest or recovery phase. The recovery phase was characterized by an increased abundance of immunoglobulins. Our work exemplifies the application of high-throughput technologies to understand adaptive processes in human physiology.

Heterotrophic carbon metabolism has been demonstrated to be limited by oxygen availability in a variety of plant tissues, which in turn inevitably affects the adenylate status. To study the effect of altering adenylate energy metabolism, without changing the oxygen supply, we expressed a plastidially targeted ATP/ADP hydrolyzing phosphatase (apyrase) in tubers of growing potato (Solanum tuberosum) plants under the control of either inducible or constitutive promoters. Inducible apyrase expression in potato tubers, for a period of 24 h, resulted in a decrease in the ATP-content and the ATP-ADP ratio in the tubers. As revealed by metabolic profiling, this was accompanied by a decrease in the intermediates of sucrose to starch conversion and several plastidially synthesized amino acids, indicating a general depression of tuber metabolism. Constitutive tuber-specific apyrase expression did not lead to a reduction of ATP, but rather a decrease in ADP and an increase in AMP levels. Starch accumulation was strongly inhibited and shifted to the production of amylopectin instead of amylose in these tubers. Furthermore, the levels of almost all amino acids were decreased, although soluble sugars and hexose-Ps were highly abundant. Respiration was elevated in the constitutively expressing lines indicating a compensation for the dramatic increase in ATP hydrolysis. The increase in respiration did not affect the internal oxygen tensions in the tubers. However, the tubers developed a ginger-like phenotype having an elevated surface-volume ratio and a reduced mass per tuber. Decreased posttranslational redox activation of ADP-glucose pyrophosphorylase and a shift in the ratio of soluble starch synthase activity to granule-bound starch synthase activity were found to be partially responsible for the alterations in starch structure and abundance. The activity of alcohol dehydrogenase was decreased and pyruvate decarboxylase was induced, but this was neither reflected by an increase in fermentation products nor in the cellular redox state, indicating that fermentation was not yet induced in the transgenic lines. When taken together the combined results of these studies allow the identification of both short- and long-term adaptation of plant metabolism and development to direct changes in the adenylate status.

Extracellular ATP (eATP) has recently been demonstrated to play a crucial role in plant development and growth. To investigate the fate of eATP within the apoplast, we used intact potato (Solanum tuberosum) tuber slices as an experimental system enabling access to the apoplast without interference of cytosolic contamination. (i) Incubation of intact tuber slices with ATP led to the formation of ADP, AMP, adenosine, adenine and ribose, indicating operation of apyrase, 5'-nucleotidase and nucleosidase. (ii) Measurement of apyrase, 5'-nucleotidase and nucleosidase activities in fractionated tuber tissue confirmed the apoplastic localization for apyrase and phosphatase in potato and led to the identification of a novel cell wall-bound adenosine nucleosidase activity. (iii) When intact tuber slices were incubated with saturating concentrations of adenosine, the conversion of adenosine into adenine was much higher than adenosine import into the cell, suggesting a potential bypass of adenosine import. Consistent with this, import of radiolabeled adenine into tuber slices was inhibited when ATP, ADP or AMP were added to the slices. (iv) In wild-type plants, apyrase and adenosine nucleosidase activities were found to be co-regulated, indicating functional linkage of these enzymes in a shared pathway. (v) Moreover, adenosine nucleosidase activity was reduced in transgenic lines with strongly reduced apoplastic apyrase activity. When taken together, these results suggest that a complete ATP salvage pathway is present in the apoplast of plant cells.

COVID-19-induced ‘acute respiratory distress syndrome’ (ARDS) is associated with prolonged respiratory failure and high mortality, but the mechanistic basis of lung injury remains incompletely understood. Here, we analyzed pulmonary immune responses and lung pathology in two cohorts of patients with COVID-19 ARDS using functional single cell genomics, immunohistology and electron microscopy. We describe an accumulation of CD163-expressing monocyte-derived macrophages that acquired a profibrotic transcriptional phenotype during COVID-19 ARDS. Gene set enrichment and computational data integration revealed a significant similarity between COVID-19-associated macrophages and profibrotic macrophage populations identified in idiopathic pulmonary fibrosis. COVID-19 ARDS was associated with clinical, radiographic, histopathological, and ultrastructural hallmarks of pulmonary fibrosis. Exposure of human monocytes to SARS-CoV-2, but not Influenza A virus or viral RNA analogs, was sufficient to induce a similar profibrotic phenotype in vitro. In conclusion, we demonstrate that SARS-CoV-2 triggers profibrotic macrophage responses and pronounced fibroproliferative ARDS., SARS-CoV-2 infection, but not influenza A, triggers immunological and pathological changes in the lung that are hallmarks of pulmonary fibrosis. A subset of CD163+ macrophages are found to drive this fibroproliferative acute respiratory distress.