Rubicon (run domain Beclin-1-interacting and cysteine-rich domain-containing protein) is a

protein Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residue (biochemistry), residues. Proteins perform a vast array of functions within organisms, including Enzyme catalysis, catalysing metab ...

that in humans is encoded by the RUBCN gene. Rubicon is one of the few known negative regulators of

autophagy Autophagy (or autophagocytosis; from the Greek language, Greek , , meaning "self-devouring" and , , meaning "hollow") is the natural, conserved degradation of the cell that removes unnecessary or dysfunctional components through a lysosome-depe ...

, a cellular process that degrades unnecessary or damaged cellular components. Rubicon is recruited to its sites of action through interaction with the

small GTPase Small GTPases (), also known as small G-proteins, are a family of hydrolase enzymes that can bind and hydrolyze guanosine triphosphate (GTP). They are a type of G-protein found in the cytosol that are homologous to the alpha subunit of heterotrim ...

Rab7 Ras-related protein Rab-7a is a protein that in humans is encoded by the ''RAB7A'' gene. Ras-related protein Rab-7a is involved in endocytosis, which is a process that brings substances into a cell. The process of endocytosis works by folding the ...

, and impairs the

autophagosome An autophagosome is a spherical structure with double layer membranes. It is the key structure in macroautophagy, the intracellular degradation system for cytoplasmic contents (e.g., abnormal intracellular proteins, excess or damaged organelles, i ...

lysosome A lysosome () is a membrane-bound organelle that is found in all mammalian cells, with the exception of red blood cells (erythrocytes). There are normally hundreds of lysosomes in the cytosol, where they function as the cell’s degradation cent ...

fusion step of autophagy through inhibition of PI3KC3-C2 (class III phosphatidylinositol 3-kinase complex 2). Negative modulation of Rubicon is associated with reduction of

aging Ageing (or aging in American English) is the process of becoming Old age, older until death. The term refers mainly to humans, many other animals, and fungi; whereas for example, bacteria, perennial plants and some simple animals are potentiall ...

and

aging-associated diseases An aging-associated disease (commonly termed age-related disease, ARD) is a disease that is most often seen with increasing frequency with increasing senescence. They are essentially complications of senescence, distinguished from the aging proce ...

: knockout of Rubicon increases lifespan in roundworms and female fruit flies, and in mice decreases kidney fibrosis and

α-synuclein Alpha-synuclein (aSyn) is a protein that in humans is encoded by the ''SNCA'' gene. It is a neuronal protein involved in the regulation of synaptic vesicle trafficking and the release of neurotransmitters. Alpha-synuclein is abundant in the bra ...

accumulation. In addition to regulation of autophagy, Rubicon has been shown to be required for LC3-associated phagocytosis (LAP) and LC3-associated endocytosis (LANDO). Rubicon has also been shown to negatively regulate the

innate immune response The innate immune system or nonspecific immune system is one of the two main immunity strategies in vertebrates (the other being the adaptive immune system). The innate immune system is an alternate defense strategy and is the dominant immune s ...

through direct interaction with multiple downstream regulatory molecules.

Structure

Rubicon consists of 972

amino acids Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although over 500 amino acids exist in nature, by far the most important are the Proteinogenic amino acid, 22 α-amino acids incorporated into p ...

and has an N-terminal

RUN domain The RUN domain is an evolutionary conserved protein–protein binding protein domain. They often interact with GTPases and could play a role in multiple Ras-like GTPase signalling pathways. This domain is present in several proteins that ar ...

, a middle region (MR), and a C-terminal Rubicon homology (RH) domain. The Rubicon homology domain is rich in cysteine residues and binds at least 4 divalent Zinc ions, forming zinc finger motifs. The structural basis for interaction between Rubicon and GTP-bound Rab7 has been experimentally determined
PDB ID: 6WCW
.

Function

The function of the N-terminal RUN domain are unknown, but it is required for autophagy suppression. The middle region contains the

PI3K Phosphoinositide 3-kinases (PI3Ks), also called phosphatidylinositol 3-kinases, are a family of enzymes involved in cellular functions such as cell growth, proliferation, differentiation, motility, survival and intracellular trafficking, which i ...

-binding domain (PIKBD), which mediates inhibition of PI3KC3-C2. The C-terminal Rubicon homology domain mediates interaction with Rab7, and is shared by other RH domain-containing autophagy regulatory proteins, including

PLEKHM1 Pleckstrin homology domain-containing family M member 1 also known as PLEKHM1 is a protein that in humans is encoded by the ''PLEKHM1'' gene. Function PLEKHM1 may have critical function in vesicular transport in osteoclasts. PLEKHM1 contain ...

and Pacer (also known as RUBCNL, Rubicon-like Autophagy Enhancer).

Autophagy-dependent

Rubicon suppresses autophagy through association with and inhibition of PI3KC3-C2. Specifically, Rubicon directly binds PI3KC3-C2 and inhibits recruitment of PI3KC3-C2 to the membrane through conformational modulation of the Beclin-1 subunit. This activity prevents PI3KC3-directed generation of

phosphatidylinositol 3-phosphate Phosphatidylinositol 3-phosphate (PI3P) is a phospholipid found in cell membranes that helps to recruit a range of proteins, many of which are involved in protein trafficking, to the membranes. It is the product of both the class II and III phosph ...

(PI3P) at the autophagosome membrane, and a resulting failure to recruit machinery that directs autophagosome-lysosome fusion. Rubicon is targeted to its site of action through direct interaction with Rab7, which decorates late endosomes and late autophagosomes.

Autophagy-independent

Rubicon has been shown to suppress the innate immune response and in some cases exacerbate viral replication. Rubicon suppresses

cytokine Cytokines () are a broad and loose category of small proteins (~5–25 kDa) important in cell signaling. Cytokines are produced by a broad range of cells, including immune cells like macrophages, B cell, B lymphocytes, T cell, T lymphocytes ...

responses through interaction with NF-κB essential modulator (NEMO), interferon regulatory factor 3 (IRF3) and caspase recruitment domain-containing protein 9 (CARD9).

Role in aging and disease

Aging-related diseases

Rubicon expression levels increase with age in mice and other model organisms, suggesting that Rubicon may cause age-associated decrease of autophagy. Since reduced autophagy is associated with aging and age-related diseases, modulation of Rubicon has been identified as a potential therapeutic target. In mice, Rubicon knockout reduces

accumulation in the brain and reduces interstitial fibrosis in the kidney.

Aging

Rubicon knockout increases lifespan in roundworms ( ''C. elegans'') through modulation of autophagy, and also increases lifespan in female fruit flies ( ''D. melanogaster'').

Nonalcoholic fatty liver disease (NAFLD)

Rubicon levels are increased in mouse models of nonalcoholic fatty liver disease (NAFLD). Knockout of Rubicon in

hepatocytes A hepatocyte is a cell of the main parenchymal tissue of the liver. Hepatocytes make up 80% of the liver's mass. These cells are involved in: * Protein synthesis * Protein storage * Transformation of carbohydrates * Synthesis of cholesterol, bile ...

improves liver steatosis and autophagy, suggesting that Rubicon contributes to NAFLD pathogenesis.

Metabolic disease

Age-dependent decline of Rubicon expression in adipose tissues may exacerbate metabolic disorders due to excessive autophagic activity.

Salih ataxia (SCAR15)

A single nucleotide deletion mutation within Rubicon is the cause of Salih ataxia
OMIM ID: 615705
. Salih ataxia (also known as spinocerebellar ataxia, autosomal recessive 15 or SCAR15) is a form of

spinocerebellar ataxia Spinocerebellar ataxia (SCA) is a progressive, degenerative, genetic disease with multiple types, each of which could be considered a neurological condition in its own right. An estimated 150,000 people in the United States have a diagnosis of ...

characterized by progressive loss of coordination of hands, gait, speech, and eye movement. The disease was discovered in children carrying a mutation (c.2624delC p.Ala875ValfsX146) causing a

frameshift mutation A frameshift mutation (also called a framing error or a reading frame shift) is a genetic mutation caused by indels ( insertions or deletions) of a number of nucleotides in a DNA sequence that is not divisible by three. Due to the triplet natur ...

and an erroneous

open reading frame In molecular biology, reading frames are defined as spans of DNA sequence between the start and stop codons. Usually, this is considered within a studied region of a prokaryotic DNA sequence, where only one of the six possible reading frames ...

in the Rubicon-coding gene starting from Alanine 875. The resulting disruption of the

C-terminal domain The C-terminus (also known as the carboxyl-terminus, carboxy-terminus, C-terminal tail, carboxy tail, C-terminal end, or COOH-terminus) is the end of an amino acid chain (protein or polypeptide), terminated by a free carboxyl group (-COOH). When t ...

impairs Rubicon subcellular localization with Rab7 and late endosomes.

References

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