[2] In 1933, while studying virgin sea urchin eggs, Jean Brachet suggested that DNA is found in cell nucleus and that RNA is present exclusively in the cytoplasm.
[3][4] During the 1930s, Joachim Hämmerling conducted experiments with Acetabularia in which he began to distinguish the contributions of the nucleus and the cytoplasm substances (later discovered to be DNA and mRNA, respectively) to cell morphogenesis and development.
This problem was overcome in the 1960s by the use of reticulocytes in vertebrates,[7] which produce large quantities of mRNA that are highly enriched in RNA encoding alpha- and beta-globin (the two major protein chains of hemoglobin).
[9] In the 1950s, results of labeling experiments in rat liver showed that radioactive amino acids were found to be associated with "microsomes" (later redefined as ribosomes) very rapidly after administration, and before they became widely incorporated into cellular proteins.
Ribosomes were first visualized using electron microscopy, and their ribonucleoprotein components were identified by biophysical methods, chiefly sedimentation analysis within ultracentrifuges capable of generating very high accelerations (equivalent to hundreds of thousands times gravity).
[11] Biochemical fractionation experiments showed that radioactive amino acids were rapidly incorporated into small RNA molecules that remained soluble under conditions where larger RNA-containing particles would precipitate.
Subsequent studies showed that (i) every cell has multiple species of tRNA, each of which is associated with a single specific amino acid, (ii) that there are a matching set of enzymes responsible for linking tRNAs with the correct amino acids, and (iii) that tRNA anticodon sequences form a specific decoding interaction with mRNA codons.
The ability to work out the genetic code emerged from the convergence of three different areas of study: (i) new methods to generate synthetic RNA molecules of defined composition to serve as artificial mRNAs, (ii) development of in vitro translation systems that could be used to translate the synthetic mRNAs into protein, and (iii) experimental and theoretical genetic work which established that the code was written in three letter "words" (codons).
Since the discovery of the tRNA cloverleaf, comparative analysis of numerous other homologous RNA molecules has led to the identification of common sequences and folding patterns.
[16] The 3569 nucleotide sequence of all of the genes of the RNA bacteriophage MS2 was determined by a large team of researchers over several years, and was reported in a series of scientific papers.
For example, mutations in HIV-1 that lead to the emergence of viral mutants that are insensitive to antiviral drugs are common, and constitute a major clinical challenge.
[20] Molecular analysis of mRNA molecules showed that, following transcription, mRNAs have non-DNA-encoded nucleotides added to both their 5′ and 3′ ends (guanosine caps and poly-A, respectively).
Key features of tRNA tertiary structure include the coaxial stacking of adjacent helices and non-Watson-Crick interactions among nucleotides within the apical loops.
In many cases, these introns were shown to be processed in more than one pattern, thus generating a family of related mRNAs that differ, for example, by the inclusion or exclusion of particular exons.
The result of alternative splicing is that a single gene can encode a number of different protein isoforms that can exhibit a variety of (usually related) biological functions.
Subsequent biochemical analysis shows that this group I intron was self-splicing; that is, the precursor RNA is capable of carrying out the complete splicing reaction in the absence of proteins.
In separate work, the RNA component of the bacterial enzyme ribonuclease P (a ribonucleoprotein complex) was shown to catalyze its tRNA-processing reaction in the absence of proteins.
[28] Introns are removed from nuclear pre-mRNAs by spliceosomes, large ribonucleoprotein complexes made up of snRNA and protein molecules whose composition and molecular interactions change during the course of the RNA splicing reactions.
[32] For years, scientists had worked to identify which protein(s) within the ribosome were responsible for peptidyl transferase function during translation, because the covalent linking of amino acids represents one of the most central chemical reactions in all of biology.
[33] Experimental methods were invented that allowed investigators to use large, diverse populations of RNA molecules to carry out in vitro molecular experiments that utilized powerful selective replication strategies used by geneticists, and which amount to evolution in the test tube.
[34] Transposable genetic elements (transposons) are found which can replicate via transcription into an RNA intermediate which is subsequently converted to DNA by reverse transcriptase.
[35] Segments of RNA, typically embedded within the 5′-untranslated region of a vast number of bacterial mRNA molecules, have a profound effect on gene expression through a previously-undiscovered mechanism that does not involve the participation of proteins.
[37] In addition to their well-established roles in translation and splicing, members of noncoding RNA (ncRNA) families have recently been found to function in genome defense and chromosome inactivation.
For example, piwi-interacting RNAs (piRNAs) prevent genome instability in germ line cells, while Xist (X-inactive-specific-transcript) is essential for X-chromosome inactivation in mammals.