NeuN (Fox-3, Rbfox3, or Hexaribonucleotide Binding Protein-3), a protein which is a homologue to the protein product of a sex-determining gene in Caenorhabditis elegans , is a neuronal nuclear antigen that is commonly used as a biomarker for neurons.
NeuN was first described in 1992 by Mullen et al., who raised a series of monoclonal antibodies to mouse antigens with the original intent of finding mouse species specific immunological markers for use in transplantation experiments. [1] In the event they isolated a hybridoma line which produced a monoclonal antibody called mAb A60, which proved to bind an antigen expressed only in neuronal nuclei and to a lesser extent the cytoplasm of neuronal cells, and which appeared to work on all vertebrates. This antigen was therefore known as NeuN for "Neuronal Nuclei" though what the A60 antibody was binding to was unknown for the next 17 years. Eventually Kim et al. used proteomic methods to show that NeuN corresponds to a protein known as Fox-3, also known as Rbfox3, a mammalian homologue of Fox-1, a protein originally identified from genetic studies of sex determination in the nematode worm Caenorhabditis elegans , see below. [2]
Western blotting shows that mAb A60 binds to two bands of apparent molecular weight ~46kDa and ~48kDa on SDS-PAGE. These two bands are generated from a single Fox-3 gene by alternate splicing. There are in fact four protein products from the Fox-3 gene as a result of the presence or absence of two amino acid sequences coded by two exons. [3] The inclusion or absence of 47 amino acids from exon 12 results in the ~46kDa and ~48kDa bands seen on SDS-PAGE gels, while the inclusion or absence of 14 amino acids from exon 15 produces two forms which are too similar in molecular size to be discerned on typical SDS-PAGE gels. Interestingly, the protein coded by exon 15 adds a C-terminal PY type nuclear localization sequence, which presumably explains why NeuN/Fox-3 protein can be both nuclear and, in some cell types, also cytoplasmic. All forms are expressed only in neurons so the mAb A60 antibody and other similar antibodies to NeuN/Fox-3 have become very widely used as robust markers of neurons.
NeuN antibodies are widely used to label neurons, despite some shortcomings, and a June 2024 Pubmed search using the keyword "NeuN" produced over 5000 hits. [4] A few neuronal cell types are not recognized by NeuN antibodies, such as Purkinje cells, stellate and Golgi cells of the cerebellum, olfactory Mitral cells, retinal photoreceptors and spinal cord gamma motor neurons. However the vast majority of neurons are strongly NeuN positive, and NeuN immunoreactivity has been widely used to identify neurons in tissue culture, in sections and to measure the neuron/glia ratio in brain regions. [5] NeuN immunoreactivity becomes obvious as neurons mature, typically after they have downregulated expression of Doublecortin, a marker seen in the earliest stages of neuronal development.
Fox-3 is one of a family of three mammalian homologues of the Fox-1 protein, originally discovered as the protein product of a gene involved in sex determination in the nematode worm C. elegans . [6] "Fox" is an acronym of "Feminizing locus on X". The mammalian genome contains three genes homologous to C. elegans Fox-1 (a.k.a. RBFOX1, A2BP, A2BP1 and HRNBP1), Fox-2, (a.k.a RBFOX2, RBM9, RTA and HRNBP2) and Fox-3 (a.k.a. RBFOX3 and HRNBP3). The HGNC names for the three proteins are RBFOX1, RBFOX2 and RBFOX3 respectively. The Fox proteins are each about 46kDa in size, and each includes a central, highly conserved ~70 amino acid RRM or RNA recognition motif. RRM domains are one of the most common in the human genome and are found in numerous proteins which bind RNA. The three mammalian Fox proteins function in the regulation of mRNA splicing and bind specific RNA sequences. [7] An alternate name the three proteins is hexaribonucleotide binding protein 1, 2 and 3, (HRNBP1, 2 and 3), since all three bind the hexaribonucleotide UGCAUG, this binding being involved in their regulation of mRNA splicing.
Alternative splicing, or alternative RNA splicing, or differential splicing, is an alternative splicing process during gene expression that allows a single gene to produce different splice variants. For example, some exons of a gene may be included within or excluded from the final RNA product of the gene. This means the exons are joined in different combinations, leading to different splice variants. In the case of protein-coding genes, the proteins translated from these splice variants may contain differences in their amino acid sequence and in their biological functions.
SR proteins are a conserved family of proteins involved in RNA splicing. SR proteins are named because they contain a protein domain with long repeats of serine and arginine amino acid residues, whose standard abbreviations are "S" and "R" respectively. SR proteins are ~200-600 amino acids in length and composed of two domains, the RNA recognition motif (RRM) region and the RS domain. SR proteins are more commonly found in the nucleus than the cytoplasm, but several SR proteins are known to shuttle between the nucleus and the cytoplasm.
Peripherin is a type III intermediate filament protein expressed mainly in neurons of the peripheral nervous system. It is also found in neurons of the central nervous system that have projections toward peripheral structures, such as spinal motor neurons. Its size, structure, and sequence/location of protein motifs is similar to other type III intermediate filament proteins such as desmin, vimentin and glial fibrillary acidic protein. Like these proteins, peripherin can self-assemble to form homopolymeric filamentous networks, but it can also heteropolymerize with neurofilaments in several neuronal types. This protein in humans is encoded by the PRPH gene. Peripherin is thought to play a role in neurite elongation during development and axonal regeneration after injury, but its exact function is unknown. It is also associated with some of the major neuropathologies that characterize amyotropic lateral sclerosis (ALS), but despite extensive research into how neurofilaments and peripherin contribute to ALS, their role in this disease is still unidentified.
Tropomyosin is a two-stranded alpha-helical, coiled coil protein found in many animal and fungal cells. In animals, it is an important component of the muscular system which works in conjunction with troponin to regulate muscle contraction. It is present in smooth and striated muscle tissues, which can be found in various organs and body systems, including the heart, blood vessels, respiratory system, and digestive system. In fungi, tropomyosin is found in cell walls and helps maintain the structural integrity of cells.
Mitochondrial 5-demethoxyubiquinone hydroxylase, also known as coenzyme Q7, hydroxylase, is an enzyme that in humans is encoded by the COQ7 gene. The clk-1 (clock-1) gene encodes this protein that is necessary for ubiquinone biosynthesis in the worm Caenorhabditis elegans and other eukaryotes. The mouse version of the gene is called mclk-1 and the human, fruit fly and yeast homolog COQ7.
RNA-binding proteins are proteins that bind to the double or single stranded RNA in cells and participate in forming ribonucleoprotein complexes. RBPs contain various structural motifs, such as RNA recognition motif (RRM), dsRNA binding domain, zinc finger and others. They are cytoplasmic and nuclear proteins. However, since most mature RNA is exported from the nucleus relatively quickly, most RBPs in the nucleus exist as complexes of protein and pre-mRNA called heterogeneous ribonucleoprotein particles (hnRNPs). RBPs have crucial roles in various cellular processes such as: cellular function, transport and localization. They especially play a major role in post-transcriptional control of RNAs, such as: splicing, polyadenylation, mRNA stabilization, mRNA localization and translation. Eukaryotic cells express diverse RBPs with unique RNA-binding activity and protein–protein interaction. According to the Eukaryotic RBP Database (EuRBPDB), there are 2961 genes encoding RBPs in humans. During evolution, the diversity of RBPs greatly increased with the increase in the number of introns. Diversity enabled eukaryotic cells to utilize RNA exons in various arrangements, giving rise to a unique RNP (ribonucleoprotein) for each RNA. Although RBPs have a crucial role in post-transcriptional regulation in gene expression, relatively few RBPs have been studied systematically.It has now become clear that RNA–RBP interactions play important roles in many biological processes among organisms.
MAFA is a type II membrane glycoprotein, first identified on the surface of rat mucosal-type mast cells of the RBL-2H3 line. More recently, human and mouse homologues of MAFA have been discovered yet also expressed by NK and T-cells. MAFA is closely linked with the type 1 Fcɛ receptors in not only mucosal mast cells of humans and mice but also in the serosal mast cells of these same organisms.
Gideon Dreyfuss is an American biochemist, the Isaac Norris Professor of Biochemistry and Biophysics at the University of Pennsylvania School of Medicine, and an investigator of the Howard Hughes Medical Institute. He was elected to the National Academy of Sciences in 2012.
Low-density lipoprotein receptor-related protein 8 (LRP8), also known as apolipoprotein E receptor 2 (ApoER2), is a protein that in humans is encoded by the LRP8 gene. ApoER2 is a cell surface receptor that is part of the low-density lipoprotein receptor family. These receptors function in signal transduction and endocytosis of specific ligands. Through interactions with one of its ligands, reelin, ApoER2 plays an important role in embryonic neuronal migration and postnatal long-term potentiation. Another LDL family receptor, VLDLR, also interacts with reelin, and together these two receptors influence brain development and function. Decreased expression of ApoER2 is associated with certain neurological diseases.
Dystonin(DST), also known as bullous pemphigoid antigen 1 (BPAG1), isoforms 1/2/3/4/5/8, is a protein that in humans is encoded by the DST gene.
RNA-binding protein with serine-rich domain 1 is a protein that in humans is encoded by the RNPS1 gene.
Regulator of nonsense transcripts 2 is a protein that in humans is encoded by the UPF2 gene.
TAR DNA-binding protein 43 is a protein that in humans is encoded by the TARDBP gene.
Regulator of nonsense transcripts 3B is a protein that in humans is encoded by the UPF3B gene.
Regulator of nonsense transcripts 3A is a protein that in humans is encoded by the UPF3A gene.
RNA binding motif protein 9 (RBM9), also known as Rbfox2, is a protein which in humans is encoded by the RBM9 gene.
Fox-1 homolog A, also known as ataxin 2-binding protein 1 (A2BP1) or hexaribonucleotide-binding protein 1 (HRNBP1) or RNA binding protein, fox-1 homolog (Rbfox1), is a protein that in humans is encoded by the RBFOX1 gene.
Messenger RNP is mRNA with bound proteins. mRNA does not exist "naked" in vivo but is always bound by various proteins while being synthesized, spliced, exported, and translated in the cytoplasm.
A neuronal lineage marker is an endogenous tag that is expressed in different cells along neurogenesis and differentiated cells such as neurons. It allows detection and identification of cells by using different techniques. A neuronal lineage marker can be either DNA, mRNA or RNA expressed in a cell of interest. It can also be a protein tag, as a partial protein, a protein or an epitope that discriminates between different cell types or different states of a common cell. An ideal marker is specific to a given cell type in normal conditions and/or during injury. Cell markers are very valuable tools for examining the function of cells in normal conditions as well as during disease. The discovery of various proteins specific to certain cells led to the production of cell-type-specific antibodies that have been used to identify cells.
Prp8 refers to both the Prp8 protein and Prp8 gene. Prp8's name originates from its involvement in pre-mRNA processing. The Prp8 protein is a large, highly conserved, and unique protein that resides in the catalytic core of the spliceosome and has been found to have a central role in molecular rearrangements that occur there. Prp8 protein is a major central component of the catalytic core in the spliceosome, and the spliceosome is responsible for splicing of precursor mRNA that contains introns and exons. Unexpressed introns are removed by the spliceosome complex in order to create a more concise mRNA transcript. Splicing is just one of many different post-transcriptional modifications that mRNA must undergo before translation. Prp8 has also been hypothesized to be a cofactor in RNA catalysis.
As of 15 June 2011, this article is derived in whole or in part from EnCor Biotechnology Inc. . The copyright holder has licensed the content in a manner that permits reuse under CC BY-SA 3.0 and GFDL. All relevant terms must be followed.The original text was at "Monoclonal Antibody to Fox3/NeuN"