Let-7 microRNA precursor
Encyclopedia
The Let-7 microRNA precursor was identified from a study of developmental timing in C. elegans
, and was later shown to be part of a much larger class of non-coding RNAs termed microRNAs. miR-98 microRNA precursor from human is a let-7 family member. Let-7 miRNAs have now been predicted or experimentally confirmed in a wide range of species (MIPF000002). miRNAs are transcribed as pri-miRNAs, which are processed in the nucleus by Drosha
and Pasha
to hairpin structures of about ~70 nucleotide
called pre-miRNAs. These precursors are exported to the cytoplasm by exportin5
, where they are subsequently processed by the enzyme Dicer
to a ~22 nucleotide mature miRNA. The involvement of Dicer in miRNA processing demonstrates a relationship with the phenomenon of RNA interference
.
) region, between loci D11S1345-D11S1316, contains the cluster miR-125b1/let-7a-2/miR-100. The cluster miR-99a/let-7c/miR-125b-2 is in a 21p11.1 region of HD (homozygous deletions). The cluster let-7g/miR-135-1 is in region 3 at 3p21.1-p21.2.
(basic local alignment search tool) research. The mature form of let-7 family members is highly conserved across species.
pathway, sequentially controlling developmental timing of larva transitions. Most animals with loss-of-function let-7 mutation burst through their vulvas and die, and therefore the mutant is lethal (let). The mutants of other let-7 family members have a radio-resistant phenotype in vulval cells, which may be related to their ability to repress RAS
.
formation in the abdomen and cell-cycle in the wing. Furthermore, the expression of pri-, pre- and mature let-7 have the same rhythmic pattern with the hormone pulse before each cuticular molt in Drosophila.
precursors of let-7 are present in these cells. It indicates that the mature let-7 miRNAs may be regulated in a post-transcriptional
manner.
cells reprogramming, LIN28
expression is reciprocal to that of mature let-7. LIN28 selectively binds the primary and precursor forms of let-7, and inhibits the processing of pri-let-7 to form the hairpin precursor. This binding is facilitated by the conserved loop sequence of primary let-7 family members and RNA-binding domains of LIN28 proteins. On the other hand, let-7 miRNAs in mammals have been shown to regulate LIN28, which implies that let-7 might enhance its own level by repressing LIN28, its negative regulator.
binding to their promoters. The levels of let-7 have been reported to decrease in models of MYC-mediated tumorigenesis, and to increase when MYC is inhibited by chemicals. In a twist, there are let-7-binding sites in MYC 3' untranslated region(UTR) according to bioinformatic analysis, and let-7 overexpression in cell culture decreased MYC mRNA levels. Therefore, there is a double-negative feedback loop between MYC and let-7. Furthermore, let-7 could lead to IMP1(/insulin-like growth factor II mRNA-binding protein) depletion, which destabilizes MYC mRNA, thus forming an indirect regulatory pathway.
), has also been identified as a target of let-7. Let-7 directly inhibits HMGA2 by binding to its 3'UTR. Removal of let-7 binding site by 3'UTR deletion cause overexpression of HMGA2 and formation of tumor. MYC is also considered as a oncogenic target of let-7.
analyses revealed many genes regulating cell cycle and cell proliferation that are responsive to alteration of let-7 levels, including cyclin A2
, CDC34
, Aurora A and B kinases
(STK6 and STK12
), E2F5
, and CDK8, among others. Subsequent experiments confirmed the direct effects of some of these genes, such as CDC25A
and CDK6. Let-7 also inhibits several components of DNA replication machinery, transcription factor
s, even some tumor suppressor gene
s and checkpoint
regulators. Apoptosis
is regulated by let-7 as well, through Casp3
, Bcl2, Map3k1
and Cdk5
modulation.
, typically in cancers that underexpress let-7. Lung cancer, for instance, have several key oncogenic mutations including p53
, RAS and MYC, part of which may directly correlates with the reduced expression of let-7, and may be repressed by introduction of let-7. Intranasal administration of let-7 has already be found effective in reducing tumor growth in a transgenic mouse model of lung cancer. Similar restoration of let-7 was also shown to inhibit cell proliferation in breast, colon and hepatic cancers, lymphoma
, and uterine leiomyoma
.
Caenorhabditis elegans
Caenorhabditis elegans is a free-living, transparent nematode , about 1 mm in length, which lives in temperate soil environments. Research into the molecular and developmental biology of C. elegans was begun in 1974 by Sydney Brenner and it has since been used extensively as a model...
, and was later shown to be part of a much larger class of non-coding RNAs termed microRNAs. miR-98 microRNA precursor from human is a let-7 family member. Let-7 miRNAs have now been predicted or experimentally confirmed in a wide range of species (MIPF000002). miRNAs are transcribed as pri-miRNAs, which are processed in the nucleus by Drosha
Drosha
Drosha is a Class 2 RNase III enzyme responsible for initiating the processing of microRNA , or short RNA molecules naturally expressed by the cell that regulate a wide variety of other genes by interacting with the RNA-induced silencing complex to induce cleavage of complementary messenger RNA ...
and Pasha
Pasha (protein)
Pasha , also known as DGCR8 in vertebrates organisms, is a protein localized to the cell nucleus that is required for microRNA processing. It binds to Drosha, an RNase III enzyme, to form the Microprocessor complex that cleaves a primary transcript known as pri-miRNA to a characteristic stem-loop...
to hairpin structures of about ~70 nucleotide
Nucleotide
Nucleotides are molecules that, when joined together, make up the structural units of RNA and DNA. In addition, nucleotides participate in cellular signaling , and are incorporated into important cofactors of enzymatic reactions...
called pre-miRNAs. These precursors are exported to the cytoplasm by exportin5
XPO5
Exportin-5 is a protein that in humans is encoded by the XPO5 gene.-Interactions:XPO5 has been shown to interact with ILF3 and Ran .-Further reading:...
, where they are subsequently processed by the enzyme Dicer
Dicer
Dicer is an endoribonuclease in the RNase III family that cleaves double-stranded RNA and pre-microRNA into short double-stranded RNA fragments called small interfering RNA about 20-25 nucleotides long, usually with a two-base overhang on the 3' end...
to a ~22 nucleotide mature miRNA. The involvement of Dicer in miRNA processing demonstrates a relationship with the phenomenon of RNA interference
RNA interference
RNA interference is a process within living cells that moderates the activity of their genes. Historically, it was known by other names, including co-suppression, post transcriptional gene silencing , and quelling. Only after these apparently unrelated processes were fully understood did it become...
.
Genomic Locations
In human genome, the cluster let-7a-1/let-7f-1/let-7d is inside the region B at 9q22.3, with the defining marker D9S280-D9S1809. One minimal LOH (loss of heterozygosityLoss of heterozygosity
Loss of heterozygosity in a cell is the loss of normal function of one allele of a gene in which the other allele was already inactivated. This term is mostly used in the context of oncogenesis; after an inactivating mutation in one allele of a tumor suppressor gene occurs in the parent's germline...
) region, between loci D11S1345-D11S1316, contains the cluster miR-125b1/let-7a-2/miR-100. The cluster miR-99a/let-7c/miR-125b-2 is in a 21p11.1 region of HD (homozygous deletions). The cluster let-7g/miR-135-1 is in region 3 at 3p21.1-p21.2.
The let-7 family
The lethal-7 (let-7) gene was first discovered in the nematode as a key developmental regulator and became one of the first two known microRNAs (the other one is lin-4). Soon, let-7 was found in fruit fly, and identified as the first known human miRNA by a BLASTBLAST
In bioinformatics, Basic Local Alignment Search Tool, or BLAST, is an algorithm for comparing primary biological sequence information, such as the amino-acid sequences of different proteins or the nucleotides of DNA sequences...
(basic local alignment search tool) research. The mature form of let-7 family members is highly conserved across species.
In C.elegans
In C.elegans, the let-7 family consists of genes encoding nine miRNAs sharing the same seed sequence. Among them, let-7, mir-84, mir-48 and mir-241 are involved in C.elegans heterochronicHeterochrony
In biology, heterochrony is defined as a developmental change in the timing of events, leading to changes in size and shape. There are two main components, namely the onset and offset of a particular process, and the rate at which the process operates...
pathway, sequentially controlling developmental timing of larva transitions. Most animals with loss-of-function let-7 mutation burst through their vulvas and die, and therefore the mutant is lethal (let). The mutants of other let-7 family members have a radio-resistant phenotype in vulval cells, which may be related to their ability to repress RAS
Ras superfamily
The Ras superfamily is a protein superfamily of small GTPases, which are all related, to a degree, to the Ras protein subfamily .There are more than a hundred proteins in the Ras superfamily...
.
In Drosophila
There is only one single let-7 gene in the Drosophila genome, which has the identical mature sequence to the one in C.elegans. The role of let-7 has been demonstrated in regulating the timing of neuromuscular junctionNeuromuscular junction
A neuromuscular junction is the synapse or junction of the axon terminal of a motor neuron with the motor end plate, the highly-excitable region of muscle fiber plasma membrane responsible for initiation of action potentials across the muscle's surface, ultimately causing the muscle to contract...
formation in the abdomen and cell-cycle in the wing. Furthermore, the expression of pri-, pre- and mature let-7 have the same rhythmic pattern with the hormone pulse before each cuticular molt in Drosophila.
In vertebrates
The let-7 family has a lot more members in vertebrates than in C.elegans and Drosophila. And the sequences, expression timing, as well as genomic clustering of these miRNAs members are all conserved across species. The direct role of let-7 family in vertebrate development has not been clearly shown as in less complex organisms, yet the expression pattern of let-7 family is indeed temporal during developmental processes. Given that the expression levels of let-7 members are significantly low in human cancers and cancer stem cells, the major function of let-7 genes may be to promote terminal differentiation in development and tumor suppression.Regulation of expression
Although the levels of mature let-7 members are undetectable in undifferentiated cells, the primary transcripts and the hairpinStem-loop
Stem-loop intramolecular base pairing is a pattern that can occur in single-stranded DNA or, more commonly, in RNA. The structure is also known as a hairpin or hairpin loop. It occurs when two regions of the same strand, usually complementary in nucleotide sequence when read in opposite directions,...
precursors of let-7 are present in these cells. It indicates that the mature let-7 miRNAs may be regulated in a post-transcriptional
Post-transcriptional regulation
Post-transcriptional regulation is the control of gene expression at the RNA level, therefore between the transcription and the translation of the gene...
manner.
By pluripotency promoting factor LIN28
As one of the four genes involved in induced pluripotent stem (iPS)Induced pluripotent stem cell
Induced pluripotent stem cells, commonly abbreviated as iPS cells or iPSCs are a type of pluripotent stem cell artificially derived from a non-pluripotent cell, typically an adult somatic cell, by inducing a "forced" expression of specific genes....
cells reprogramming, LIN28
LIN28
Lin-28 homolog A is a protein that in humans is encoded by the LIN28 gene. It is a marker of undifferentiated human embryonic stem cells and has been used to enhance the efficiency of the formation of induced pluripotent stem cells from human fibroblasts...
expression is reciprocal to that of mature let-7. LIN28 selectively binds the primary and precursor forms of let-7, and inhibits the processing of pri-let-7 to form the hairpin precursor. This binding is facilitated by the conserved loop sequence of primary let-7 family members and RNA-binding domains of LIN28 proteins. On the other hand, let-7 miRNAs in mammals have been shown to regulate LIN28, which implies that let-7 might enhance its own level by repressing LIN28, its negative regulator.
In autoregulatory loop with MYC
Expression of let-7 members is controlled by MYCMyc
Myc is a regulator gene that codes for a transcription factor. In the human genome, Myc is located on chromosome 8 and is believed to regulate expression of 15% of all genes through binding on Enhancer Box sequences and recruiting histone acetyltransferases...
binding to their promoters. The levels of let-7 have been reported to decrease in models of MYC-mediated tumorigenesis, and to increase when MYC is inhibited by chemicals. In a twist, there are let-7-binding sites in MYC 3' untranslated region(UTR) according to bioinformatic analysis, and let-7 overexpression in cell culture decreased MYC mRNA levels. Therefore, there is a double-negative feedback loop between MYC and let-7. Furthermore, let-7 could lead to IMP1(/insulin-like growth factor II mRNA-binding protein) depletion, which destabilizes MYC mRNA, thus forming an indirect regulatory pathway.
Oncogenes: RAS, HMGA2
Let-7 has been demonstrated to be a direct regulator of RAS expression in human cellsRAS genes in human, K-, N-, and H-, have the predicted let-7 binding sequences in their 3'UTRs. In lung cancer patient samples, expression of RAS and let-7 showed reciprocal pattern, which has low let-7 and high RAS in cancerous cells, and high let-7 and low RAS in normal cells. Another oncogene, high mobility group A2 (HMGA2HMGA2
High-mobility group AT-hook 2, also known as HMGA2, is a protein that, in humans, is encoded by the HMGA2 gene.- Function :This gene encodes a protein that belongs to the non-histone chromosomal high-mobility group protein family. HMG proteins function as architectural factors and are essential...
), has also been identified as a target of let-7. Let-7 directly inhibits HMGA2 by binding to its 3'UTR. Removal of let-7 binding site by 3'UTR deletion cause overexpression of HMGA2 and formation of tumor. MYC is also considered as a oncogenic target of let-7.
Cell cycle, proliferation, and apoptosis regulators
MicroarrayMicroarray
A microarray is a multiplex lab-on-a-chip. It is a 2D array on a solid substrate that assays large amounts of biological material using high-throughput screening methods.Types of microarrays include:...
analyses revealed many genes regulating cell cycle and cell proliferation that are responsive to alteration of let-7 levels, including cyclin A2
Cyclin A2
Cyclin-A2 is a protein that in humans is encoded by the CCNA2 gene.-Interactions:Cyclin A2 has been shown to interact with ITGB3BP, Retinoblastoma-like protein 1, E2F1, CDC6, SKP2 and Flap structure-specific endonuclease 1.-References:...
, CDC34
CDC34
Ubiquitin-conjugating enzyme E2 R1 is a protein that in humans is encoded by the CDC34 gene.-Interactions:CDC34 has been shown to interact with CSNK2B, BTRC and CDK9.-Further reading:...
, Aurora A and B kinases
Aurora Kinase
Aurora kinases are serine/threonine kinases that are essential for cell proliferation. The enzyme helps the dividing cell dispense its genetic materials to its daughter cells. More specifically, Aurora kinases play a crucial role in cellular division by controlling chromatid segregation...
(STK6 and STK12
Aurora B kinase
Aurora B kinase is a protein that functions in the attachment of the mitotic spindle to the centromere.In cancerous cells, over-expression of these enzymes causes unequal distribution of genetic information, creating aneuploid cells, a hallmark of cancer....
), E2F5
E2F5
Transcription factor E2F5 is a protein that in humans is encoded by the E2F5 gene.-Further reading:...
, and CDK8, among others. Subsequent experiments confirmed the direct effects of some of these genes, such as CDC25A
CDC25A
Cell division cycle 25 homolog A , also known as CDC25A, is a mammalian gene.-Interactions:CDC25A has been shown to interact with ASK1, Epidermal growth factor receptor, C-Raf, CHEK1, Cyclin E1, PIM1 and YWHAB.-Further reading:...
and CDK6. Let-7 also inhibits several components of DNA replication machinery, transcription factor
Transcription factor
In molecular biology and genetics, a transcription factor is a protein that binds to specific DNA sequences, thereby controlling the flow of genetic information from DNA to mRNA...
s, even some tumor suppressor gene
Tumor suppressor gene
A tumor suppressor gene, or anti-oncogene, is a gene that protects a cell from one step on the path to cancer. When this gene is mutated to cause a loss or reduction in its function, the cell can progress to cancer, usually in combination with other genetic changes.-Two-hit hypothesis:Unlike...
s and checkpoint
Cell cycle checkpoint
Cell cycle checkpoints are control mechanisms that ensure the fidelity of cell division in eukaryotic cells. These checkpoints verify whether the processes at each phase of the cell cycle have been accurately completed before progression into the next phase...
regulators. Apoptosis
Apoptosis
Apoptosis is the process of programmed cell death that may occur in multicellular organisms. Biochemical events lead to characteristic cell changes and death. These changes include blebbing, cell shrinkage, nuclear fragmentation, chromatin condensation, and chromosomal DNA fragmentation...
is regulated by let-7 as well, through Casp3
Caspase 3
Caspase 3 is a caspase protein that interacts with caspase 8 and caspase 9. It is encoded by the CASP3 gene. CASP3 orthologs have been identified in numerous mammals for which complete genome data are available...
, Bcl2, Map3k1
MAP3K1
Mitogen-activated protein kinase kinase kinase 1 is an enzyme that in humans is encoded by the MAP3K1 gene.Mouse genetics has revealed that the kinase is important in: correct embryogenesis, keratinocyte migration, T cell cytokine production and B cell antibody production.-Interactions:MAP3K1 has...
and Cdk5
Cyclin-dependent kinase 5
Cell division protein kinase 5 is an enzyme that in humans is encoded by the CDK5 gene. The protein encoded by this gene is part of the cyclin-dependent kinase family.-Physiological Role:...
modulation.
Potential clinical use in cancer
Given the prominent phenotype of cell overproliferation and undifferentiation by let-7 loss-of-function in nematodes, and the role of its targets on cell destiny determination, let-7 is closely associated with human cancer and acts as a tumor suppressor.Diagnosis
Numerous reports have shown that the expression levels of let-7 are frequently low and the chromosomal clusters of let-7 are often deleted in many cancers. Let-7 is expressed at higher levels in more differentiated tumors, which also have lower levels of activated oncogenes such as RAS and HMGA2. Therefore, expression levels of let-7 could be prognostic markers in several cancers associated with differentiation stages. In lung cancer, for example, reduced expression of let-7 is significantly correlated with reduced postoperative survival.Therapy
Let-7 is also a very attractive potential therapeutic that can prevent tumorigenesis and angiogenesisAngiogenesis
Angiogenesis is the physiological process involving the growth of new blood vessels from pre-existing vessels. Though there has been some debate over terminology, vasculogenesis is the term used for spontaneous blood-vessel formation, and intussusception is the term for the formation of new blood...
, typically in cancers that underexpress let-7. Lung cancer, for instance, have several key oncogenic mutations including p53
P53
p53 , is a tumor suppressor protein that in humans is encoded by the TP53 gene. p53 is crucial in multicellular organisms, where it regulates the cell cycle and, thus, functions as a tumor suppressor that is involved in preventing cancer...
, RAS and MYC, part of which may directly correlates with the reduced expression of let-7, and may be repressed by introduction of let-7. Intranasal administration of let-7 has already be found effective in reducing tumor growth in a transgenic mouse model of lung cancer. Similar restoration of let-7 was also shown to inhibit cell proliferation in breast, colon and hepatic cancers, lymphoma
Lymphoma
Lymphoma is a cancer in the lymphatic cells of the immune system. Typically, lymphomas present as a solid tumor of lymphoid cells. Treatment might involve chemotherapy and in some cases radiotherapy and/or bone marrow transplantation, and can be curable depending on the histology, type, and stage...
, and uterine leiomyoma
Leiomyoma
A leiomyoma is a benign smooth muscle neoplasm that is not premalignant. They can occur in any organ, but the most common forms occur in the uterus, small bowel and the esophagus.- Etymology:* Greek:** λεῖος leios "smooth"...
.