1Pooja Mankad and2Ankit V. Kachchhi

For a considerable period, the prevailing belief was that RNA primarily served as a messenger, conveying instructions from the genes in DNA to the proteins responsible for cellular functions. In addition to this role, specific types of RNA, such as tRNA and rRNA, played roles in cell maintenance. However, the landscape shifted significantly with the discovery of the first short non-coding RNA, known as microRNA, approximately 30 years ago. This discovery opened the field of non-coding RNAs with no or little protein-coding potential. Since then, many new classes of non-coding RNAs, including endogenous small interfering RNAs (endo-siRNAs), PIWI-associated RNAs (piRNAs), and long non-coding RNAs, have been identified. Non-coding RNA (ncRNA) refers to a class of RNA molecules that do not encode proteins but instead play essential roles in regulating gene expression and various cellular processes. These molecules contribute to the complexity of genetic control by influencing transcription, translation, and other key aspects of cellular function.

Types of ncRNAs

There are two main types of non-coding RNAs viz., Housekeeping ncRNAs and Regulatory ncRNAs.

  1. HousekeepingncRNAs: These RNAs are referred to as “housekeeping” because they are constitutively expressed in all cells and are involved in essential cellular processes necessary for the maintenance of cell structure and function. Housekeeping RNAs are typically stable and have relatively constant expression levels across different cell types and under various physiological conditions.

The most common types of housekeeping RNAs include:

  1. rRNA (ribosomal RNA): The majority of the RNA in the cell is ribosomal RNA, making up over 80 percent of the total cellular RNA. It is synthesized in the nucleolus, which lies within the nucleus of the cell, from rDNA by RNA polymerase type I. In association with ribosomal proteins, it forms the ribosomal structure. It also interacts with transfer RNA and assists in protein synthesis, thus having structural and functional roles. Prokaryotic 70S ribosomes are made up of 5S, 23S, and 16S rRNA subunits, while eukaryotic 80S ribosomal subunits include 5S, 28S, 5.8S, and 18S rRNA subunits.
  2. tRNA (transfer RNA): tRNA synthesis is directed by RNA polymerase III and occurs in the nucleus of the cell. Pre-tRNA undergoes splicing to form mature tRNA. Two major things must be remembered about tRNA: it has an anticodon arm that bind to the complementary triplet codon on the messenger RNA and decodes the message, and it has an amino acid binding arm on which a specific amino acid is bound and carried to ribosomes for protein chain elongation. Another interesting feature about tRNA is that the third anticodon base, called the wobble base, allows the same tRNA to read multiple codons on mRNA.
  3. snRNA (small nuclear RNA): They are about 150 base pairs long and are transcribed by RNA polymerase type II or type III. Their major role is to process the pre-mRNA in the nucleus. However, they are also involved in transcription factor regulation and telomere maintenance. RNA complexes with other proteins to form snRNP (small nuclear ribonucleoprotein) spliceosomal complexes that help in splicing the transcripts.
  4. snoRNA (small nucleolar RNA): They are involved in the chemical modification of other RNA molecules. These are involved in the covalent modification of rRNA, tRNA, and snRNA, mainly through methylation or pseudo-uridilation. To carry out such modifications, these RNAs guide the RNP (ribonuclear protein) complex forming snoRNP complex, which leads to covalent modifications of the transcripts.
  1. Regulatory ncRNAs: Regulatory RNAs are a class of non-coding RNAs (ncRNAs) that play crucial roles in the regulation of gene expression and other cellular processes. Unlike housekeeping RNAs, which are involved in basic cellular functions and are constitutively expressed in all cells, regulatory RNAs are more context-dependent and often show dynamic expression patterns in response to specific conditions. These RNAs participate in the control of gene expression at various levels, contributing to the complexity and precision of cellular regulation.

Several types of regulatory RNAs include:

  1. miRNA (microRNA): It is one of the first and most extensively studied regulatory non-coding RNAs. microRNA is endogenously produced as a single-stranded RNA that folds to form a stem-loop structure. miRNA originates from intronic or exonic regions in the nucleus as primary miRNA. The primary miRNA is cleaved by Drosha to form pre-miRNA, which is exported from the nucleus to the cytosol. Pre-miRNA is further cleaved by Dicer proteins, giving rise to the final single-stranded miRNA. miRNA binds to the target mRNA by complementary base pairing, and with the aid of other endonucleases or by forming the RISC (RNA-induced silencing complex), it leads to mRNA cleavage or inhibits translation
  2. siRNA (short interfering RNA):They are usually exogenous in origin or synthesized as short double-stranded RNA, similar to miRNA. They are about 20 to 25 base pairs in length, and their major role is in RNA interference technology, which leads to gene silencing through mRNA cleavage. siRNA is generally used to validate gene function by post-transcriptional cleavage or degradation of target mRNA
  3. piRNA (piwi-interacting RNA): They are also short, about 24 to 31 nucleotides long, and generally expressed from transposon regions. Their major role is in transcriptional regulation and silencing. piRNA interacts with Piwi or the piwi proteins and guides them to transposon transcripts.
  4. lncRNA (long non-coding RNA): They are generally over 200 nucleotides long andsimilar to protein-coding mRNA in many forms, such as they show splicing, polyadenylation, and other modifications. But the unique feature includes that they do not code for proteins, primarily because they lack the open reading frame (ORF), which is essential for translation. Secondly, they show several-fold lower expression compared to mRNA transcripts. One of the major challenges in the identification and prediction of such long non-coding RNAs is that they share little conservation across species. They may be involved in transcriptional or translational regulation. They may bind to other RNA molecules, such as miRNA or mRNA. They interact with other splicing factors and alter the splicing of transcripts. They may also lead to epigenetic changes by recruiting chromatin remodelling complexes, such as histone methyltransferases. They may block transcription factor binding to promoters and regulate gene expression.

Role of ncRNAs

Non-coding RNAs (ncRNAs) play diverse and crucial roles in various cellular processes, contributing significantly to the complexity of gene regulation and the maintenance of cellular homeostasis. Some key roles of non-coding RNAs include:

  1. Gene Regulation:

MicroRNAs (miRNAs) and Small Interfering RNAs (siRNAs): These small RNAs regulate gene expression by binding to target messenger RNAs (mRNAs), leading to their degradation or translational repression.

  1. Epigenetic Regulation:

Long Non-Coding RNAs (lncRNAs): Involved in chromatin remodelling and modification, influencing gene expression patterns epigenetically.

  1. Chromatin Structure and Organization:

lncRNAs: Contribute to the three-dimensional organization of chromatin, affecting the accessibility of genes to transcriptional machinery.

  1. Post-Transcriptional Modifications:

Small Nuclear RNAs (snRNAs) and Small Nucleolar RNAs (snoRNAs): Involved in pre-mRNA splicing and post-transcriptional modifications of other RNAs, such as rRNAs and tRNAs.

  1. Ribosome Biogenesis:

snoRNAs: Guide chemical modifications in ribosomal RNA during ribosome biogenesis.

  1. Transposon Silencing:

Piwi-Interacting RNAs (piRNAs): Protect the genome by silencing transposable elements, particularly in germ cells.

  1. Cellular Stress Response:

Various ncRNAs: Respond to environmental stressors and cellular insults, influencing cellular survival and adaptation.

  1. Cell Cycle Regulation:

Various ncRNAs: Contribute to the regulation of the cell cycle by influencing the expression of genes involved in cell division and growth.

  1. Immune Response:

Various ncRNAs: Modulate immune responses by regulating the expression of genes involved in immune system functions.

  1. Developmental Processes:

Various ncRNAs: Play roles in embryonic development, tissue differentiation, and organogenesis.