An overlapping gene (or OLG)
is a
gene
In biology, the word gene has two meanings. The Mendelian gene is a basic unit of heredity. The molecular gene is a sequence of nucleotides in DNA that is transcribed to produce a functional RNA. There are two types of molecular genes: protei ...
whose expressible
nucleotide sequence
A nucleic acid sequence is a succession of bases within the nucleotides forming alleles within a DNA (using GACT) or RNA (GACU) molecule. This succession is denoted by a series of a set of five different letters that indicate the order of the nu ...
partially overlaps with the expressible nucleotide sequence of another gene.
In this way, a nucleotide sequence may make a contribution to the function of one or more
gene product
A gene product is the biochemical material, either RNA or protein, resulting from the expression of a gene. A measurement of the amount of gene product is sometimes used to infer how active a gene is. Abnormal amounts of gene product can be corre ...
s. Overlapping genes are present in and a fundamental feature of both
cellular and
viral
The word ''Viral'' means "relating to viruses" (small infectious agents).
It may also refer to:
Viral behavior, or virality
Memetic behavior likened that of a virus, for example:
* Viral marketing, the use of existing social networks to spre ...
genome
A genome is all the genetic information of an organism. It consists of nucleotide sequences of DNA (or RNA in RNA viruses). The nuclear genome includes protein-coding genes and non-coding genes, other functional regions of the genome such as ...
s.
The current definition of an overlapping gene varies significantly between eukaryotes, prokaryotes, and viruses.
In
prokaryote
A prokaryote (; less commonly spelled procaryote) is a unicellular organism, single-celled organism whose cell (biology), cell lacks a cell nucleus, nucleus and other membrane-bound organelles. The word ''prokaryote'' comes from the Ancient Gree ...
s and
virus
A virus is a submicroscopic infectious agent that replicates only inside the living Cell (biology), cells of an organism. Viruses infect all life forms, from animals and plants to microorganisms, including bacteria and archaea. Viruses are ...
es overlap must be between
coding sequences but not
mRNA
In molecular biology, messenger ribonucleic acid (mRNA) is a single-stranded molecule of RNA that corresponds to the genetic sequence of a gene, and is read by a ribosome in the process of Protein biosynthesis, synthesizing a protein.
mRNA is ...
transcripts, and is defined when these coding sequences share a nucleotide on either the same or opposite strands. In
eukaryote
The eukaryotes ( ) constitute the Domain (biology), domain of Eukaryota or Eukarya, organisms whose Cell (biology), cells have a membrane-bound cell nucleus, nucleus. All animals, plants, Fungus, fungi, seaweeds, and many unicellular organisms ...
s, gene overlap is almost always defined as mRNA transcript overlap. Specifically, a gene overlap in eukaryotes is defined when at least one nucleotide is shared between the boundaries of the primary mRNA transcripts of two or more genes, such that a DNA base
mutation
In biology, a mutation is an alteration in the nucleic acid sequence of the genome of an organism, virus, or extrachromosomal DNA. Viral genomes contain either DNA or RNA. Mutations result from errors during DNA or viral replication, ...
at any point of the overlapping region would affect the transcripts of all genes involved. This definition includes 5′ and 3′
untranslated region
In molecular genetics, an untranslated region (or UTR) refers to either of two sections, one on each side of a coding sequence on a strand of mRNA. If it is found on the Directionality (molecular biology), 5' side, it is called the Five prime ...
s (UTRs) along with
intron
An intron is any nucleotide sequence within a gene that is not expressed or operative in the final RNA product. The word ''intron'' is derived from the term ''intragenic region'', i.e., a region inside a gene."The notion of the cistron .e., gen ...
s.
Overprinting refers to a type of overlap in which all or part of the sequence of one gene is read in an alternate
reading frame
In molecular biology, a reading frame is a specific choice out of the possible ways to read the nucleic acid sequence, sequence of nucleotides in a nucleic acid (DNA or RNA) molecule as a sequence of triplets. Where these triplets equate to amino ...
from another gene at the same
locus.
The alternative open reading frames (ORF) are thought to be created by critical
nucleotide substitutions within an expressible pre-existing gene, which can be
induced to express a novel
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 ...
while still preserving the function of the original gene. Overprinting has been hypothesized as a mechanism for
''de novo'' emergence of new genes from existing sequences, either older genes or previously
non-coding
Non-coding DNA (ncDNA) sequences are components of an organism's DNA that do not encode protein sequences. Some non-coding DNA is transcribed into functional non-coding RNA molecules (e.g. transfer RNA, microRNA, piRNA, ribosomal RNA, and regula ...
regions of the genome.
It is believed that most overlapping genes, or genes whose expressible nucleotide sequences partially overlap with each other, evolved in part due to this mechanism, suggesting that each overlap is composed of one ancestral gene and one novel gene. Subsequently, overprinting is also believed to be a source of novel proteins, as de novo proteins coded by these novel genes usually lack remote
homologs
Homologous chromosomes or homologs are a set of one maternal and one paternal chromosome that pair up with each other inside a cell during meiosis. Homologs have the same genes in the same loci, where they provide points along each chromosome th ...
in databases. Overprinted genes are particularly common features of the
genomic
Genomics is an interdisciplinary field of molecular biology focusing on the structure, function, evolution, mapping, and editing of genomes. A genome is an organism's complete set of DNA, including all of its genes as well as its hierarchical, ...
organization of viruses, likely to greatly increase the number of potential expressible genes from a small set of viral genetic information.
It is likely that overprinting is responsible for the generation of numerous novel proteins by viruses over the course of their
evolutionary history
The history of life on Earth traces the processes by which living and extinct organisms evolved, from the earliest emergence of life to the present day. Earth formed about 4.5 billion years ago (abbreviated as ''Ga'', for '' gigaannum'') and ...
.
Classification
Genes may overlap in a variety of ways and can be classified by their positions relative to each other.
* ''Unidirectional'' or ''tandem'' overlap: the
3' end of one gene overlaps with the
5' end of another gene on the same strand. This arrangement can be symbolized with the notation → → where arrows indicate the reading frame from start to end.
* ''Convergent'' or ''end-on'' overlap: the
3' ends of the two genes overlap on opposite strands. This can be written as → ←.
* ''Divergent'' or ''tail-on'' overlap: the
5' ends of the two genes overlap on opposite strands. This can be written as ← →.
Overlapping genes can also be classified by ''phases'', which describe their relative
reading frame
In molecular biology, a reading frame is a specific choice out of the possible ways to read the nucleic acid sequence, sequence of nucleotides in a nucleic acid (DNA or RNA) molecule as a sequence of triplets. Where these triplets equate to amino ...
s:
*''In-phase overlap'' occurs when the shared sequences use the same reading frame. This is also known as "phase 0". Unidirectional genes with phase 0 overlap are not considered distinct genes, but rather as
alternative start site
The start codon is the first codon of a messenger RNA (mRNA) transcript translated by a ribosome. The start codon always codes for methionine in eukaryotes and archaea and a ''N''-formylmethionine (fMet) in bacteria, mitochondria and plastids.
T ...
s of the same gene.
*''Out-of-phase overlaps'' occurs when the shared sequences use different reading frames. This can occur in "phase 1" or "phase 2", depending on whether the reading frames are offset by 1 or 2 nucleotides. Because a
codon
Genetic code is a set of rules used by living cells to translate information encoded within genetic material (DNA or RNA sequences of nucleotide triplets or codons) into proteins. Translation is accomplished by the ribosome, which links prote ...
is three nucleotides long, an offset of three nucleotides is an in-phase, phase 0 frame.
Studies on overlapping genes suggest that their evolution can be summarized in two possible models.
In one model, the two proteins encoded by their respective overlapping genes evolve under similar
selection pressures. The proteins and the overlap region are highly conserved when strong selection against
amino acid
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 22 α-amino acids incorporated into proteins. Only these 22 a ...
change is favored. Overlapping genes are reasoned to evolve under strict constraints as a single nucleotide substitution is able to alter the structure and function of the two proteins simultaneously. A study on the
hepatitis B virus
Hepatitis B virus (HBV) is a partially double-stranded DNA virus, a species of the genus '' Orthohepadnavirus'' and a member of the '' Hepadnaviridae'' family of viruses. This virus causes the disease hepatitis B.
Classification
Hepatitis B ...
(HBV), whose DNA genome contains numerous overlapping genes, showed the mean number of synonymous nucleotide substitutions per site in overlapping coding regions was significantly lower than that of non-overlapping regions.
The same study showed that it was possible for some of these overlapping regions and their proteins to diverge significantly from the original when there's weak selection against amino acid change. The
spacer domain of the
polymerase
In biochemistry, a polymerase is an enzyme (Enzyme Commission number, EC 2.7.7.6/7/19/48/49) that synthesizes long chains of polymers or nucleic acids. DNA polymerase and RNA polymerase are used to assemble DNA and RNA molecules, respectively, by ...
and the pre-S1 region of a surface protein of HBV, for example, had a percentage of conserved amino acids of 30% and 40%, respectively.
However, these overlap regions are known to be less important for
replication compared to the overlap regions that were highly conserved among different HBV strains, which are absolutely essential for the process.
The second model suggests that the two proteins and their respective overlap genes evolve under opposite selection pressures: one frame experiences
positive selection
In population genetics, directional selection is a type of natural selection in which one extreme phenotype is favored over both the other extreme and moderate phenotypes. This genetic selection causes the allele frequency to shift toward the ...
while the other is under
purifying selection
In natural selection, negative selection or purifying selection is the selective removal of alleles that are deleterious. This can result in stabilising selection through the purging of deleterious genetic polymorphisms that arise through random ...
. In
tombusvirus
''Tombusvirus'' is a genus of viruses, in the family ''Tombusviridae''. Plants serve as natural hosts. There are 17 species in this genus. Symptoms associated with this genus include mosaic. The name of the genus comes from Tomato bushy stunt v ...
es, the proteins
p19 and
p22 are encoded by overlapping genes that form a 549 nt coding region, and p19 is shown to be under positive selection while p22 is under purifying selection. Additional examples are mentioned in studies involving overlapping genes of the
Sendai virus
''Murine respirovirus'', formerly ''Sendai virus'' (SeV) and previously also known as murine parainfluenza virus type 1 or hemagglutinating virus of Japan (HVJ), is an Viral envelope, enveloped, 150-200 nm–diameter, negative sense, single ...
,
potato leafroll virus
Potato leafroll virus (PLRV) is a member of the genus '' Polerovirus'' and family '' Solemoviridae''. The phloem limited positive sense RNA virus infects potatoes and other members of the family Solanaceae. PLRV was first described by Quanjer ...
, and human
parvovirus B19
Parvovirus B19, also called B19 virus (B19V), Human parvovirus B19, or sometimes erythrovirus B19, is a human virus in the family ''Parvoviridae'', genus ''Erythroparvovirus''. It measures only 23–26 nm in diameter. The virus is assigned ...
. This phenomenon of overlapping genes experiencing different selection pressures is suggested to be a consequence of a high
rate of nucleotide substitution with different effects on the two frames; the substitutions may be majorly
non-synonymous for one frame while mostly being
synonymous
A synonym is a word, morpheme, or phrase that means precisely or nearly the same as another word, morpheme, or phrase in a given language. For example, in the English language, the words ''begin'', ''start'', ''commence'', and ''initiate'' are a ...
for the other frame.
Evolution
Overlapping genes are particularly common in rapidly evolving genomes, such as those of
virus
A virus is a submicroscopic infectious agent that replicates only inside the living Cell (biology), cells of an organism. Viruses infect all life forms, from animals and plants to microorganisms, including bacteria and archaea. Viruses are ...
es,
bacteria
Bacteria (; : bacterium) are ubiquitous, mostly free-living organisms often consisting of one Cell (biology), biological cell. They constitute a large domain (biology), domain of Prokaryote, prokaryotic microorganisms. Typically a few micr ...
, and
mitochondria
A mitochondrion () is an organelle found in the cells of most eukaryotes, such as animals, plants and fungi. Mitochondria have a double membrane structure and use aerobic respiration to generate adenosine triphosphate (ATP), which is us ...
. They may originate in three ways:
# By extension of an existing
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 ...
(ORF) downstream into a contiguous gene due to the loss of a
stop codon
In molecular biology, a stop codon (or termination codon) is a codon (nucleotide triplet within messenger RNA) that signals the termination of the translation process of the current protein. Most codons in messenger RNA correspond to the additio ...
;
# By extension of an existing ORF upstream into a contiguous gene due to loss of an
initiation codon
The start codon is the first codon of a messenger RNA (mRNA) transcript translated by a ribosome. The start codon always codes for methionine in eukaryotes and archaea and a ''N''-formylmethionine (fMet) in bacteria, mitochondria and plastids.
T ...
;
# By generation of a novel ORF within an existing one due to a
point mutation
A point mutation is a genetic mutation where a single nucleotide base is changed, inserted or deleted from a DNA or RNA sequence of an organism's genome. Point mutations have a variety of effects on the downstream protein product—consequences ...
.
The use of the same nucleotide sequence to encode multiple genes may provide
evolution
Evolution is the change in the heritable Phenotypic trait, characteristics of biological populations over successive generations. It occurs when evolutionary processes such as natural selection and genetic drift act on genetic variation, re ...
ary advantage due to reduction in
genome
A genome is all the genetic information of an organism. It consists of nucleotide sequences of DNA (or RNA in RNA viruses). The nuclear genome includes protein-coding genes and non-coding genes, other functional regions of the genome such as ...
size and due to the opportunity for
transcriptional
Transcription is the process of copying a segment of DNA into RNA for the purpose of gene expression. Some segments of DNA are transcribed into RNA molecules that can encode proteins, called messenger RNA (mRNA). Other segments of DNA are transc ...
and
translational co-regulation
Co-regulation (or coregulation) is a term used in psychology. It is defined most broadly as a "continuous unfolding of individual action that is susceptible to being continuously modified by the continuously changing actions of the partner". An imp ...
of the overlapping genes.
Gene overlaps introduce novel evolutionary constraints on the sequences of the overlap regions.
Origins of new genes

In 1977,
Pierre-Paul Grassé
Pierre-Paul Grassé (November 27, 1895 in Périgueux (Dordogne) – July 9, 1985) was a French zoologist, writer of over 300 publications including the influential 52-volume '' Traité de Zoologie''. He was an expert on termites who rejected Neo- ...
proposed that one of the genes in the pair could have originated ''de novo'' by mutations to introduce novel ORFs in alternate reading frames; he described the mechanism as ''overprinting''.
It was later substantiated by
Susumu Ohno
was a Japanese-American geneticist and evolutionary biologist, and seminal researcher in the field of molecular evolution.
Biography
Susumu Ohno was born to Japanese parents in Keijō, Chōsen (present-day Seoul, South Korea), Empire of ...
, who identified a candidate gene that may have arisen by this mechanism.
Some de novo genes originating in this way may not remain overlapping, but
subfunctionalize following
gene duplication
Gene duplication (or chromosomal duplication or gene amplification) is a major mechanism through which new genetic material is generated during molecular evolution. It can be defined as any duplication of a region of DNA that contains a gene ...
,
contributing to the prevalence of
orphan gene
Orphan genes, ORFans, or taxonomically restricted genes (TRGs) are genes that lack a detectable homologue outside of a given species or lineage. Most genes have known homologues. Two genes are homologous when they share an evolutionary history, a ...
s. Which member of an overlapping gene pair is younger can be identified
bioinformatic
Bioinformatics () is an interdisciplinary field of science
Science is a systematic discipline that builds and organises knowledge in the form of testable hypotheses and predictions about the universe. Modern science is typically divi ...
ally either by a more restricted
phylogenetic
In biology, phylogenetics () is the study of the evolutionary history of life using observable characteristics of organisms (or genes), which is known as phylogenetic inference. It infers the relationship among organisms based on empirical dat ...
distribution, or by less optimized
codon usage.
Younger members of the pair tend to have higher
intrinsic structural disorder than older members, but the older members are also more disordered than other proteins, presumably as a way of alleviating the increased evolutionary constraints posed by overlap.
Overlaps are more likely to originate in proteins that already have high disorder.
Taxonomic distribution
Overlapping genes occur in all
domains of life, though with varying frequencies. They are especially common in
viral
The word ''Viral'' means "relating to viruses" (small infectious agents).
It may also refer to:
Viral behavior, or virality
Memetic behavior likened that of a virus, for example:
* Viral marketing, the use of existing social networks to spre ...
genomes.
Viruses
The existence of overlapping genes was first identified in the virus
ΦX174
The phi X 174 (or ΦX174) bacteriophage is a single-stranded DNA ( ssDNA) virus that infects ''Escherichia coli''. This virus was isolated in 1935 by Nicolas Bulgakov in Félix d'Hérelle's laboratory at the Pasteur Institute, from samples co ...
, whose
genome
A genome is all the genetic information of an organism. It consists of nucleotide sequences of DNA (or RNA in RNA viruses). The nuclear genome includes protein-coding genes and non-coding genes, other functional regions of the genome such as ...
was the first DNA genome ever sequenced by
Frederick Sanger
Frederick Sanger (; 13 August 1918 – 19 November 2013) was a British biochemist who received the Nobel Prize in Chemistry twice.
He won the 1958 Chemistry Prize for determining the amino acid sequence of insulin and numerous other prote ...
in 1977.
Previous analysis of ΦX174, a small single-stranded DNA
bacteriophage
A bacteriophage (), also known informally as a phage (), is a virus that infects and replicates within bacteria. The term is derived . Bacteriophages are composed of proteins that Capsid, encapsulate a DNA or RNA genome, and may have structu ...
that infected the bacteria
Escherichia coli
''Escherichia coli'' ( )Wells, J. C. (2000) Longman Pronunciation Dictionary. Harlow ngland Pearson Education Ltd. is a gram-negative, facultative anaerobic, rod-shaped, coliform bacterium of the genus '' Escherichia'' that is commonly fo ...
, suggested that the
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 ...
s produced during infection required
coding sequences longer than the measured length of its genome.
Analysis of the fully sequenced 5386 nucleotide genome showed that the virus possessed extensive overlap between coding regions, revealing that some genes (like genes D and E) were translated from the same DNA sequences but in different reading frames.
An
alternative start site
The start codon is the first codon of a messenger RNA (mRNA) transcript translated by a ribosome. The start codon always codes for methionine in eukaryotes and archaea and a ''N''-formylmethionine (fMet) in bacteria, mitochondria and plastids.
T ...
within the genome replication gene A of ΦX174 was shown to express a
truncated protein with an identical coding sequence to the
C-terminus
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
Proteins are large biomolecules and macromolecules that comp ...
of the original A protein but possessing a different function It was concluded that other undiscovered sites of
polypeptide synthesis
Protein biosynthesis, or protein synthesis, is a core biological process, occurring inside cells, balancing the loss of cellular proteins (via degradation or export) through the production of new proteins. Proteins perform a number of critical ...
could be hidden through the genome due to overlapping genes. An identified de novo gene of another overlapping
gene locus
In genetics, a locus (: loci) is a specific, fixed position on a chromosome where a particular gene or genetic marker is located. Each chromosome carries many genes, with each gene occupying a different position or locus; in humans, the total numb ...
was shown to express a novel protein that induces lysis of E. coli by inhibiting biosynthesis of its cell wall
6 suggesting that de novo protein creation through the process of overprinting can be a significant factor in the evolution of
pathogenicity
In biology, a pathogen (, "suffering", "passion" and , "producer of"), in the oldest and broadest sense, is any organism or agent that can produce disease. A pathogen may also be referred to as an infectious agent, or simply a germ.
The term ...
of viruses.
Another example is the ''
ORF3d
ORF3d is a gene found in SARS-CoV-2 (the virus that causes COVID-19) and at least one closely related coronavirus found in pangolins, though it is not found in other closely related viruses within the '' Sarbecovirus'' subgenus. It is 57 codons l ...
'' gene in the
SARS-CoV 2 virus.
Overlapping genes are particularly common in
viral
The word ''Viral'' means "relating to viruses" (small infectious agents).
It may also refer to:
Viral behavior, or virality
Memetic behavior likened that of a virus, for example:
* Viral marketing, the use of existing social networks to spre ...
genomes.
Some studies attribute this observation to
selective pressure
Evolutionary pressure, selective pressure or selection pressure is exerted by factors that reduce or increase reproductive success in a portion of a population, driving natural selection. It is a quantitative description of the amount of change oc ...
toward small genome sizes mediated by the physical constraints of packaging the genome in a
viral capsid
A capsid is the protein shell of a virus, enclosing its genetic material. It consists of several oligomeric (repeating) structural subunits made of protein called protomers. The observable 3-dimensional morphological subunits, which may or ma ...
, particularly one of
icosahedral
In geometry, an icosahedron ( or ) is a polyhedron with 20 faces. The name comes . The plural can be either "icosahedra" () or "icosahedrons".
There are infinitely many non- similar shapes of icosahedra, some of them being more symmetrical tha ...
geometry.
However, other studies dispute this conclusion and argue that the distribution of overlaps in viral genomes is more likely to reflect overprinting as the evolutionary origin of overlapping viral genes.
Overprinting is a common source of ''de novo'' genes in viruses.
The proportion of viruses with overlapping coding sequences within their genomes varies.
Double-stranded
RNA virus
An RNA virus is a virus characterized by a ribonucleic acid (RNA) based genome. The genome can be single-stranded RNA (ssRNA) or double-stranded (Double-stranded RNA, dsRNA). Notable human diseases caused by RNA viruses include influenza, SARS, ...
es have fewer than a quarter that contains them while almost three-quarters of
retroviridae
A retrovirus is a type of virus that inserts a DNA copy of its RNA genome into the DNA of a host cell that it invades, thus changing the genome of that cell. After invading a host cell's cytoplasm, the virus uses its own reverse transcriptase ...
and viruses with
single-stranded DNA genomes contain overlapping coding sequences.
Segmented viruses in particular, or viruses with their genome split into separate pieces and packaged either all in the same
capsid
A capsid is the protein shell of a virus, enclosing its genetic material. It consists of several oligomeric (repeating) structural subunits made of protein called protomers. The observable 3-dimensional morphological subunits, which may or m ...
or in separate capsids, are more likely to contain an overlapping sequence than non-segmented viruses.
RNA viruses have fewer overlapping genes than DNA viruses which possess lower
mutation rate
In genetics, the mutation rate is the frequency of new mutations in a single gene, nucleotide sequence, or organism over time. Mutation rates are not constant and are not limited to a single type of mutation; there are many different types of mu ...
s and less restrictive genome sizes.
The lower mutation rate of DNA viruses facilitates greater genomic novelty and evolutionary exploration within a structurally constrained genome and may be the primary driver of the evolution of overlapping genes.
Studies of overprinted viral genes suggest that their protein products tend to be accessory proteins which are not
essential to viral proliferation, but contribute to
pathogenicity
In biology, a pathogen (, "suffering", "passion" and , "producer of"), in the oldest and broadest sense, is any organism or agent that can produce disease. A pathogen may also be referred to as an infectious agent, or simply a germ.
The term ...
. Overprinted proteins often have unusual
amino acid
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 22 α-amino acids incorporated into proteins. Only these 22 a ...
distributions and high levels of intrinsic
disorder
Disorder may refer to randomness, a lack of intelligible pattern, or:
Healthcare
* Disorder (medicine), a functional abnormality or disturbance
* Mental disorder or psychological disorder, a psychological pattern associated with distress or disab ...
.
In some cases overprinted proteins do have well-defined, but novel, three-dimensional structures;
one example is the
RNA silencing suppressor p19 found in
Tombusvirus
''Tombusvirus'' is a genus of viruses, in the family ''Tombusviridae''. Plants serve as natural hosts. There are 17 species in this genus. Symptoms associated with this genus include mosaic. The name of the genus comes from Tomato bushy stunt v ...
es, which has both a novel
protein fold
A protein superfamily is the largest grouping (clade) of proteins for which common ancestry can be inferred (see homology). Usually this common ancestry is inferred from structural alignment and mechanistic similarity, even if no sequence similar ...
and a novel binding mode in recognizing
siRNA
Small interfering RNA (siRNA), sometimes known as short interfering RNA or silencing RNA, is a class of double-stranded non-coding RNA molecules, typically 20–24 base pairs in length, similar to microRNA (miRNA), and operating within the RN ...
s.
Prokaryotes
Estimates of gene overlap in
bacteria
Bacteria (; : bacterium) are ubiquitous, mostly free-living organisms often consisting of one Cell (biology), biological cell. They constitute a large domain (biology), domain of Prokaryote, prokaryotic microorganisms. Typically a few micr ...
l genomes typically find that around one third of bacterial genes are overlapped, though usually only by a few base pairs.
Most studies of overlap in bacterial genomes find evidence that overlap serves a function in
gene regulation
Regulation of gene expression, or gene regulation, includes a wide range of mechanisms that are used by cells to increase or decrease the production of specific gene products (protein or RNA). Sophisticated programs of gene expression are wide ...
, permitting the overlapped genes to be
transcriptionally and
translationally co-regulated.
In prokaryotic genomes, unidirectional overlaps are most common, possibly due to the tendency of adjacent prokaryotic genes to share orientation.
Among unidirectional overlaps, long overlaps are more commonly read with a one-nucleotide offset in reading frame (i.e., phase 1) and short overlaps are more commonly read in phase 2.
Long overlaps of greater than 60
base pair
A base pair (bp) is a fundamental unit of double-stranded nucleic acids consisting of two nucleobases bound to each other by hydrogen bonds. They form the building blocks of the DNA double helix and contribute to the folded structure of both DNA ...
s are more common for convergent genes; however, putative long overlaps have very high rates of
misannotation.
Robustly validated examples of long overlaps in bacterial genomes are rare; in the well-studied
model organism
A model organism is a non-human species that is extensively studied to understand particular biological phenomena, with the expectation that discoveries made in the model organism will provide insight into the workings of other organisms. Mo ...
''
Escherichia coli
''Escherichia coli'' ( )Wells, J. C. (2000) Longman Pronunciation Dictionary. Harlow ngland Pearson Education Ltd. is a gram-negative, facultative anaerobic, rod-shaped, coliform bacterium of the genus '' Escherichia'' that is commonly fo ...
'', only four gene pairs are well validated as having long, overprinted overlaps.
Eukaryotes
Compared to prokaryotic genomes, eukaryotic genomes are often poorly annotated and thus identifying genuine overlaps is relatively challenging.
However, examples of validated gene overlaps have been documented in a variety of eukaryotic organisms, including mammals such as mice and humans.
Eukaryotes differ from prokaryotes in distribution of overlap types: while unidirectional (i.e., same-strand) overlaps are most common in prokaryotes, opposite or antiparallel-strand overlaps are more common in eukaryotes. Among the opposite-strand overlaps, convergent orientation is most common.
Most studies of eukaryotic gene overlap have found that overlapping genes are extensively subject to genomic reorganization even in closely related species, and thus the presence of an overlap is not always well-conserved.
Overlap with older or less taxonomically restricted genes is also a common feature of genes likely to have originated ''de novo'' in a given eukaryotic lineage.
Function
The precise functions of overlapping genes seems to vary across the domains of life but several experiments have shown that they are important for virus lifecycles through proper protein expression and stoichiometry as well as playing a role in proper protein folding. A version of
bacteriophage
A bacteriophage (), also known informally as a phage (), is a virus that infects and replicates within bacteria. The term is derived . Bacteriophages are composed of proteins that Capsid, encapsulate a DNA or RNA genome, and may have structu ...
ΦX174
The phi X 174 (or ΦX174) bacteriophage is a single-stranded DNA ( ssDNA) virus that infects ''Escherichia coli''. This virus was isolated in 1935 by Nicolas Bulgakov in Félix d'Hérelle's laboratory at the Pasteur Institute, from samples co ...
has also been created where all gene overlaps were removed proving they were not necessary for replication.
The retention and evolution of overlapping genes within viruses may also be due to
capsid
A capsid is the protein shell of a virus, enclosing its genetic material. It consists of several oligomeric (repeating) structural subunits made of protein called protomers. The observable 3-dimensional morphological subunits, which may or m ...
size limitations. Dramatic viability loss was observed in viruses with genomes engineered to be longer than the wild-type genome. Increasing the single-stranded DNA genome length of
ΦX174
The phi X 174 (or ΦX174) bacteriophage is a single-stranded DNA ( ssDNA) virus that infects ''Escherichia coli''. This virus was isolated in 1935 by Nicolas Bulgakov in Félix d'Hérelle's laboratory at the Pasteur Institute, from samples co ...
by >1% results in almost complete loss of
infectivity
In epidemiology, infectivity is the ability of a pathogen
In biology, a pathogen (, "suffering", "passion" and , "producer of"), in the oldest and broadest sense, is any organism or agent that can produce disease. A pathogen may also be refer ...
, believed to be the result of the strict physical constraints imposed by the finite capsid volume. Studies on
adeno-associated virus
Adeno-associated viruses (AAV) are small viruses that infect humans and some other primate species. They belong to the genus '' Dependoparvovirus'', which in turn belongs to the family ''Parvoviridae''. They are small (approximately 26 nm in ...
es as
gene delivery
Gene delivery is the process of introducing foreign genetic material, such as DNA or RNA, into host cells. Gene delivery must reach the genome of the host cell to induce gene expression. Successful gene delivery requires the foreign gene deliver ...
vectors showed that viral packaging is constrained by genetic cargo size limits, requiring the use of multiple
vectors to deliver large human genes such as CFTR81. Therefore, it is suggested that overlapping genes evolved as a means to overcome these physical constraints, increasing genetic diversity by utilizing only the existing sequence rather than increasing genome length.
Methods in identifying overlapping genes and ORFs
Standardized methods such as
genome annotation
In molecular biology and genetics, DNA annotation or genome annotation is the process of describing the structure and function of the components of a genome, by analyzing and interpreting them in order to extract their biological significance and ...
may be inappropriate for the detection of overlapping genes as they are reliant on already curated genes while overlapping genes are generally overlooked contain atypical sequence composition.
Genome annotation standards are also often biased against feature overlaps, such as genes entirely contained within another gene. Furthermore, some bioinformatics pipelines such as the
RAST pipeline markedly penalizes overlaps between predicted ORFs. However, rapid advancement of genome-scale protein and RNA measurement tools along with increasingly advanced prediction algorithms have revealed an avalanche of overlapping genes and ORFs within numerous genomes.
Proteogenomic methods have been essential in discovering numerous overlapping genes and include a combination of techniques such as
bottom-up proteomics
Bottom-up proteomics is a common method to identify proteins and characterize their amino acid sequences and post-translational modifications by proteolytic digestion of proteins prior to analysis by mass spectrometry.
BUP techniques can be an a ...
,
ribosome profiling
Ribosome profiling, or Ribo-Seq (also named ribosome footprinting), is an adaptation of a technique developed by Joan Steitz and Marilyn Kozak almost 50 years ago that Nicholas Ingolia and Jonathan Weissman adapted to work with next generation se ...
,
DNA sequencing
DNA sequencing is the process of determining the nucleic acid sequence – the order of nucleotides in DNA. It includes any method or technology that is used to determine the order of the four bases: adenine, thymine, cytosine, and guanine. The ...
, and Perturb-seq, perturbation. RNA-Seq, RNA sequencing is also used to identify genomic regions containing overlapping transcripts. It has been utilized to identify 180,000 alternate ORFs within previously annotated coding regions found in humans. Newly discovered ORFs such as these are verified using a variety of reverse genetics techniques, such as CRISPR gene editing, CRISPR-Cas9 and CRISPR activation#dCas9, catalytically dead Cas9 (dCas9) disruption. Attempts at proof-by-synthesis are also performed to show beyond doubt the absence of any undiscovered overlapping genes.
See also
* Nested gene
References
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Genes