An Autoantigen Atlas From Human Lung HFL1 Cells Offers Clues to Neurological and Diverse Autoimmune Manifestations of COVID-19

Abstract

COVID-19 is accompanied by a myriad of both transient and long-lasting autoimmune responses. Dermatan sulfate (DS), a glycosaminoglycan crucial for wound healing, has unique affinity for autoantigens (autoAgs) from apoptotic cells. DS-autoAg complexes are capable of stimulating autoreactive B cells and autoantibody production. We used DS-affinity proteomics to define the autoantigen-ome of lung fibroblasts and bioinformatics analyses to study the relationship between autoantigenic proteins and COVID-induced alterations. Using DS-affinity, we identified an autoantigen-ome of 408 proteins from human HFL1 cells, at least 231 of which are known autoAgs. Comparing with available COVID data, 352 proteins of the autoantigen-ome have thus far been found to be altered at protein or RNA levels in SARS-CoV-2 infection, 210 of which are known autoAgs. The COVID-altered proteins are significantly associated with RNA metabolism, translation, vesicles and vesicle transport, cell death, supramolecular fibrils, cytoskeleton, extracellular matrix, and interleukin signaling. They offer clues to neurological problems, fibrosis, smooth muscle dysfunction, and thrombosis. In particular, 150 altered proteins are related to the nervous system, including axon, myelin sheath, neuron projection, neuronal cell body, and olfactory bulb. An association with the melanosome is also identified. The findings from our study illustrate a connection between COVID infection and autoimmunity. The vast number of COVID-altered proteins with high intrinsic propensity to become autoAgs offers an explanation for the diverse autoimmune complications in COVID patients. The variety of autoAgs related to mRNA metabolism, translation, and vesicles suggests a need for long-term monitoring of autoimmunity in COVID. The COVID autoantigen atlas we are establishing provides a detailed molecular map for further investigation of autoimmune sequelae of the pandemic, such as "long COVID" syndrome.

Summary sentence: An autoantigen-ome by dermatan sulfate affinity from human lung HFL1 cells may explain neurological and autoimmune manifestations of COVID-19.

Keywords: COVID-19; SARS-CoV-2; autoantibodies; autoantigens; autoimmunity; dermatan sulfate.

Figure 1

The 408-protein autoantigen-ome identified by DS-affinity from HFL1 cells forms a highly interacting network. Connecting lines represent interactions with high confidence (minimum interaction score of 0.7) as per STRING analysis. Colored proteins are involved in metabolism of RNA (blue), vesicles (pink), cytoskeleton (gold), collagen and elastic fibers (light green), and chondroitin sulfate/dermatan sulfate metabolism (dark green).

Figure 2

Network of 352 autoantigen-ome proteins that are altered in SARS-CoV-2 infected cells or patients. Connecting lines represent interactions with high confidence. Colored proteins are involved in metabolism of RNA (77 proteins, red), mRNA metabolic process (69 proteins, gold), translation (43 proteins, pink), vesicles (99 proteins, light green) and vesicle-mediated transport (84 proteins, dark green), cytoskeleton (84 proteins, blue), and extracellular matrix organization (32 proteins, aqua).

Figure 3

(A) Interaction network of 260 up-regulated proteins in SARS-CoV-2 infected cells or patients. Connecting lines represent interactions with high confidence (minimum interaction score of 0.7). Colored proteins are involved in metabolism of RNA (54 proteins, red), translation (28 proteins, pink), vesicles (82 proteins, light green) and vesicle-mediated transport (67 proteins, dark green), regulation of cell death (61 proteins, blue), and mRNA metabolic process (46 proteins, gold). (B) Interaction network of 303 down-regulated proteins in SARS-Cov-2 infected cells and patients. Connecting lines represent interactions with high confidence. Marked proteins are involved in RNA metabolism (64 proteins), translation (39 proteins, pink), vesicles (88 proteins, green), cytoskeleton (73 proteins, blue), and extracellular matrix organization (29 proteins, aqua).

Figure 4

(A) Nervous system-related proteins among COVID-altered proteins. Colored proteins are involved in axon guidance (62 proteins, aqua), axon growth cone (25 proteins, blue), myelin sheath (26 proteins, red), neuron projection (32 proteins, green) and neuron projection extension (7 proteins, dark green), neuronal cell body (16 proteins, gold), peripheral nervous system axon regeneration (3 proteins, brown), cerebellar Purkinje cell layer development (4 proteins, amber), and olfactory bulb (23 proteins, pink). (B) Neurological disease-related proteins among proteins altered in COVID. Colored are proteins found in neuronal infection with Japanese encephalitis virus (23 proteins, blue), neuroblastoma (21 proteins, red), glioblastoma (22 proteins, pink), neurodegeneration in Down syndrome (26 proteins, dark green), Alzheimer disease (22 proteins, aqua), schizophrenia (24 proteins, amber), cerebral ischemia induced neurodegenerative diseases (17 proteins, dark purple), Parkinson disease (17 proteins, brown), and neurodegeneration (21 proteins, green).

Figure 5

(A) Relation of COVID-altered proteins to wound healing and hemostasis. Response to wounding (25 proteins, red), blood vessel development (20 proteins, pink), blood coagulation (14 proteins, blue), collagen-containing extracellular matrix (13 proteins, brown), collagen biosynthesis and modifying enzymes (16 proteins, dark purple), platelet activation (3 proteins, dark green) and platelet activation signaling and aggregation (22 proteins, green), platelet degranulation (18 proteins, aqua), and hemostasis (35 proteins, gold). (B) Other significantly enriched groups among altered proteins. Supramolecular fiber (56 proteins, amber), melanosome (30 proteins, brown), striated muscle cell differentiation (11 proteins, purple), myofibril (23 proteins, red), muscle structure development (18 proteins, green), muscle contraction (13 proteins, aqua), Z disk (9 proteins, dark green), intercalated disk (4 proteins, blue), and amyloid fiber formation (6 proteins, pink).

Figure 6

Interaction network of altered proteins in 6 COVID-19 patients. Colored proteins are associated with leukocyte activation involved in immune response (red), vesicles (light green) and vesicle-mediated transport (dark green), protein processing in the ER (yellow), regulation of cell death (blue), translation (pink), melanosome (brown), myelin sheath (aqua), and muscle contraction (amber).

Figure 7

(A) Hierarchical clustering of top 10 pathways involving COVID-altered proteins. Analysis based on hypergeometric distribution followed by FDR correction. (B) COVID-altered host proteins with DS-affinity found in various viral infections. Porcine reproductive and respiratory syndrome (56 proteins, green), H5N1 avian influenza virus (27 proteins, dark purple), Japanese encephalitis virus (23 proteins, gold), Rift Valley fever virus (24 proteins, aqua), Hepatitis B virus (22 proteins, dark green), HIV (identified in different studies, 18 amber, 18 brown, 18 red and 17 pink), and shared among positive-sense RNA viruses (20 proteins, blue).