Let-7 MicroRNA Precursor

The Let-7 microRNA precursor gives rise to let-7, a microRNA (miRNA) involved in control of stem-cell division and differentiation. ''let-7'', short for "lethal-7", was discovered along with the miRNA ''lin-4'' in a study of developmental timing in '' C. elegans'', making these miRNAs the first ever discovered. ''let-7'' was later identified in humans as the first human miRNA , and is highly conserved across many species. Dysregulation of ''let-7'' contributes to cancer development in humans by preventing differentiation of cells, leaving them stuck in a stem-cell like state. ''let-7'' is therefore classified as a tumor suppressor.

The ''let-7 microRNA family'' refers to the many slight variations of ''let-7'' that exist both within a single organism and across species. In humans, for example, there are ten unique ''let-7'' family member sequences: ''let-7''a through ''g'', ''let-7i'', ''mir-98'', and ''mir-202''.

In humans, mature ''let-7'' acts via RNA-induced silencing by complexing with RISC and binding to target mRNA, preventing translation into protein. Known targets of ''let''-7 include proteins related to the cell cycle and proliferation, such as MYC, RAS, cyclin D, HMGA2, and CDC25A. Knockdown of these proteins by ''let-7'' prevents proliferation and induces differentiation of cells. Important inhibitors of ''let-7'' include LIN28, which binds to ''let-7'' directly, and the proto-oncogene MYC, which suppresses expression.

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 heterozygosity) 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 ''C. elegans'' 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 the fruit fly (''Drosophila)'', and identified as the first known human miRNA by a BLAST (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 the ''C. elegans'' heterochronic 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.

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 junction 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''. 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 temporally regulated during developmental processes. Functionally, ''let-7'' has been shown in early vertebrates to control the differentiation of mesoderm and ectoderm. 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 hairpin 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.

By pluripotency promoting factor ''LIN28''

As one of the genes involved in (but not essential for) induced pluripotent stem (iPS) cell 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. Lin-28 uses two zinc knuckle domains to recognize the NGNNG motif in the ''let-7'' precursors, while the Cold-shock domain, connected by a flexible linker, binds to a closed loop in the precursors. 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 MYC 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.

Targets of ''let-7''

Oncogenes: ''RAS'', ''HMGA2''

''Let-7'' has been demonstrated to be a direct regulator of ''RAS'' expression in human cells All the three ''RAS'' 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'' ('' HMGA2''), 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.

Cell cycle, proliferation, and apoptosis regulators

Microarray 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 factors, even some tumor suppressor genes and checkpoint regulators. Apoptosis is regulated by ''let-7'' as well, through Casp3, Bcl2, Map3k1 and Cdk5 modulation.He YJ, Guo L, D ZH. (2009) Let-7 and mir-24 in uvb-induced apoptosis hinese Zhonghua Fang She Yi Xue Yu Fang Hu Za Zhi. 29, 234–6.

Immunity

''Let-7'' has been implicated in post-transcriptional control of innate immune responses to pathogenic agents. Macrophages stimulated with live bacteria or purified microbial components down-regulate the expression of several members of the ''let-7'' microRNA family to relieve repression of immune-modulatory cytokines IL-6 and IL-10. ''Let-7'' has also been implicated in the negative regulation of TLR4, the major immune receptor of microbial lipopolysaccharide and down-regulation of ''let-7'' both upon microbial and protozoan infection might elevate TLR4 signaling and expression. ''Let-7'' has furthermore been reported to regulate the production of cytokine IL-13 by T lymphocytes during allergic airway inflammation thus linking this microRNA to adaptive immunity as well. Down-modulation of ''let-7'' negative regulator '' Lin28b'' in human T lymphocytes is believed to accrue during early neonate development to reprogram the immune system towards defense.

Potential clinical use in cancer

Given the prominent phenotype of cell overproliferation and dedifferentiation 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.

Prognostic Biomarkers

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. The expression of ''let-7b'' and ''let-7g'' microRNAs are significantly associated with overall survival in 1262 breast cancer patients.

Therapy

''Let-7'' is also a very attractive potential therapeutic that can prevent tumorigenesis and angiogenesis, typically in cancers that underexpress ''let-7''. Lung cancer, for instance, has several key oncogenic mutations including ''p53'', ''RAS'' and ''MYC'', some of which may directly correlate with the reduced expression of ''let-7'', and may be repressed by introduction of ''let-7''. Intranasal administration of ''let-7'' has already been 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.

References

Further reading

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External links

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miRBase family entry for let-7

{{miRNA precursor families

MicroRNA