Кажучи “минуле в минулому”, ми жертвуємо майбутнім. Sir Winston Leonard Spencer-Churchill
Які асоціації у Вас викликає фраза: “Монстр Шпігельмана”?
Sol Spiegelman monster is a fragment of ss(+)RNA bacteriphage genome copied many times by RNA-dependent-RNA-polymerase
Two scenarios from Segré & Lancet (2000) A – RNA first (strong RNA world hypothesis) B – Lipids first (lipid world hypothesis – compositional genomes – metabolism without genes)
Are there alternatives to RNA? RNA a – Threose Nucleic Acid – TNA b – Peptide nucleic acid – PNA c – Glycerol derived nucleic acid d – Pyranosyl RNA hybridizes with other nucleic acids. Information is not lost. DNA-RNA hybrids DNA takes over at end of RNA world. Maybe TNA or PNA preceded the RNA world. Information passed to RNA. Would need to show that the alternative was easier to synthesize than RNA.
Deoxyribozymes: -RNA cleavage -DNA phosphorylation -DNA adenylation -DNA deglycosylation -Thymine dimer photoreversion -DNA cleavage -DNA ligation Ronald R. Breaker discovered deoxyribozymes in 1994 DNAzymes have low efficiency. They catalyze reactions at ~ 100 fold rate
Alexander Rich - the author of RNA world concept Walter Gilbert RNA World Lesley Orgel Francis Crick
Puzzle of RNA World
Ribose synthesis in presence of borate minerals solves the “asphalt” problem
John Sutherland in 2009 showed the possibility to synthesize pyrimidine ribonucleotides (C, T) in alternative root The starting materials for the synthesis—cyanamide, cyanoacetylene, glycolaldehyde, glyceraldehyde and inorganic phosphate—are plausible prebiotic feedstock molecules
Гіпотеза світу поліароматичних вуглеводнів (Simon Nicholas Platts, 2004)
p. H decreased
Another model of RNA polimerization is self-assembling
Next step: RNA replication Non-enzymatic templatedirected polymerization Polymerization by a general RNA polymerase ribozyme (or replicase) Mutually autocatalytic networks of replicases
Non-enzymatic templatedirected Polymerization Deck, C. , Jauker, M. & Richert, C. Efficient enzyme-free copying of all four nucleobases templated by immobilized RNA. Nature Chem. 3, 603– 608 (2011 ) It could generate products up to 30 -50 nt
RNA polymerization by a general RNA polymerase ribozyme. "Molecular biologist's dream" Joyce, Orgel RNA polymerase ribozyme B 6. 61 replicates template of >200 nt by blocks of 20 nt
Mutually autocatalytic network
Emergence of hypercycles
Transition from chemical state to biological
RNA world idea originated in 60’s as a theoretical solution to the chicken and egg problem of DNA and proteins. Self-splicing introns. First RNA catalysts to be discovered. Tom Cech (1982). ‘RNA World’ term coined by Walter Gilbert (1986).
Example of an RNA catalyst Hammerhead ribozyme Cleaves RNA at a specific point. Rolling circle mechanism of replication of virus-like RNAs in plants. Chops long strand into pieces.
What can ribozymes do? Ligases E’ T. A. Lincoln, G. F. Joyce, Science 323, 1229 (2009)
What can ribozymes do? Recombinases E. J. Hayden, G. v. Kiedrowski & N. Lehman, Angew. Chem. Int. Edit. (2008) 120, 8552 Catalyst is autocatalytic given a supply of W X Y Z. The non-covalent assembly is also a catalyst.
What can ribozymes do? Polymerases Black +Blue – ribozyme Red – template Orange – primer Primer extended by up to 14 nucleotides Johnstone et al. (2001) Science
Gradual improvement of Polymerases in the lab Wochner et al. (2011) Science - up to 95 nucleotides
What can ribozymes do? Nucleotide Synthetases Unrau and Bartel, (1998) Nature
An RNA organism must have had a metabolism. Hypothetical pathway for RNA catalyzed RNA synthesis (Joyce) Synthesis of nucleosides Phosphorylation Generation of NTPs Creation of activated nucleotides Stepwise polymerization
From RNA to DNA transition Схематичне зображення еволюції геному від РНК до модифікованих ДНК геномів. У віросфері представлені всі типи геномів, а в сучасних клітинах – лише ДНК-Т геном. Останній міг виникнути внаслідок перенесення ДНК від дволанцюгового ДНК вірусу в РНК клітину. RNR – рибонуклеотид редуктаза; Td. S – тимідилат синтетаза; Hmc. T – гідроксиметилцитозин трансфераза.
LUCA – last universal common ancestor
On the Origin of Genetic Code
Триплетність Безперервність Дискретність Специфічність Виродженість Колінеарність Універсальність Нерандомність
Інші властивості генетичного коду Залежність між другим нуклеотидом кодону і властивостями амінокислоти, що кодується; Зв’язок із кодоном в другій позиції і класом аміноацил-т. РНК синтетази; Негативна кореляція між молекулярною масою амінокислоти і кількістю кодонів, що її кодують; Позитивна кореляція між числом синонімічних кодонів для амінокислоти і частотою з якою ця амінокислота зустрічається в білках; Принцип певної мінімізації негативних ефектів точкових мутацій та помилок при трансляції
Prefix Root Ending XYZ Stem CUU CUC CUA CUG Leu GUU GUC GUA GUG NAN: Asp, Glu, Lys, Asn, Gln, His, Tyr Polar NUN: Leu, Ile, Val, Phe, Met Hydrophobic NCN: Ala, Ser, Thr, Pro Small NGN: Cys, Trp, Arg, Gly, Ser Special Val
Principal Component Analysis Projects the 8 -d space into the two ‘most important’ dimensions. Big Small Hydrophobic Hydrophilic
Anticodon, Codon, 1 st position 3 d position C C 3' C C G 5' → U U 34 36 m. I I G C G A G G G ψ → → A, G G → U, C I → U, C, A
Origin and evolution of the genetic code: the universal enigma Eugene V. Koonin* and Artem S. Novozhilov 2009
Theories Of Genetic Code Evolution Francis Crick – author of ‘frozen accident theory’ of universality of genetic code 1966 Stereochemical theory Error minimization theory Coevolution theory
Stereochemical theory
Cost function g(a, b) for replacing amino acid a by amino acid b e. g. difference in Polar Requirement rij = rate of mistaking codon i for codon j = 1 for single position mistakes, 0 otherwise E = measure of error associated with a code Generate random codes by permuting the 20 amino acids in the code table E is smaller for the canonical code than for almost all random codes. the probability of a random code to be fitter than the standard one is P 1 ≈10− 4 there are more than 1084 possible alternative codes f ~ 10 -6 p(E) Ereal one in a million codes is better (Freeland Hurst) f E
r 1, r 2 ∈ r : random codes with the same block structure as the standard code o 1, o 2 ∈ o : codes obtained from r 1, r 2 ∈ r after optimization R 1, R 2 ∈ R : random codes with fitness values greater than the fitness of the standard code O 1, O 2 ∈ O : codes obtained from R 1, R 2 ∈ R after optimization
Origin and evolution of the genetic code: the universal enigma Eugene V. Koonin* and Artem S. Novozhilov 2009
An optimized genetic code with the same block structure and degeneracy level as the standard code
The earliest code probably had few amino acids. Which were the first? Selection acts when new amino acids are added. Maybe only 2 nd position was relevant initially. U C A G U C U A G U C C Val A Late amino acids took over codons previously assigned to amino acids with similar properties. Ala Asp Gly A G U C A G Propose that the four earliest amino acids were Val, Ala, Asp, Gly
Code structure after addition of the 10 early amino acids. . Add new amino acids in positions that were formerly occupied by amino acids with similar properties. This minimizes disruption to existing gene sequences.
Pathways of amino acid synthesis in modern organisms (from Di Giulio 2008)
Variables of genetic code are examples of its evolution
Codon Reassignment – The Genetic code is variable in mitochondria (and also some cases of other types of genomes) Second Position U F i r s t P o s i t i o n U C A G Third Pos. F F L L S S Y Y Stop C C Stop W U C A G L L P P H H Q Q R R U C A G CUN Leu to Thr I I I M T T N N K K S S R R U C A G AGR Arg to Ser to Stop/Gly V V A A D D E E G G U C A G UGA Stop to Trp AUA Ile to Met CGN Arg to unassigned etc. . . But how can this happen? It should be disadvantageous.
Reassignments in Metazoa Porifera Cnidaria Arthropoda Nematoda Lophotrochozoa Loss of t. RNA-Ile(CAU) but AUA remains Ile Loss of t. RNA-Arg(UCU) and AGR : Arg -> Ser Loss of many t. RNAs + import from cytoplasm Platyhelminthes Echinodermata Hemichordata AUA : Ile -> Met AGR : Ser -> Stop Urochordata AGR : Ser -> Gly AAA : Lys -> Asn AAA : Lys -> unassigned Cephalochordata Craniata
Example 1: AUA was reassigned from Ile to Met during the early evolution of the mitochondrial genome. Before Codon Anticodon Ile Ile Met After Ile Met AUU AUC AUA GAU k 2 CAU AUG CAU Codon Anticodon AUU AUC Notes G in the wobble position of the t. RNA-Ile can pair with U and C in the third codon position Bacteria and some protist mitochondria possess another t. RNA-Ile with a modified base that translates AUA only. The t. RNA-Met translates AUG only. GAU AUA AUG Notes In animal mitochondria the k 2 CAU t. RNA has been deleted. UAU or f 5 CAU There is a gain of function of the t. RNA-Met by a mutation or a base modification
Example 2: UGA was reassigned from Stop to Trp many times (12 times in mitochondria). Before Codon Anticodon Notes Stop UGA RF Release Factor recognizes UGA codon. Trp UGG CCA Normal t. RNA-Trp translates only UGG codons. After Codon Anticodon Trp UGA UGG UCA Notes In animal mitochondria (and elsewhere) there is a gain of function of the t. RNA-Trp via mutation or base modification so that it translates both UGG and UGA.
The GAIN-LOSS framework (Sengupta & Higgs, Genetics 2005) LOSS = deletion or loss of function of a t. RNA or RF GAIN = gain of a new t. RNA or a gain of function of an existing one. GAIN Ambiguous codon. Selective disadvantage. New Code. Selective disadvantage because codons are used in wrong places Initial Code. No Problem. LOSS Unassigned codon. Selective disadvantage. Note – the strength of the selective disadvantage depends on the number of times the codon is used. There is no disadvantage if the codon disappears. GAIN Mutations in coding sequences New Code. Codons now used in right places. No Problem.
‘Genome streamlining’ - the selective pressure to minimize mitochondrial genomes yields reassignments of specific codons, in particular, one of the three stop codons.
Modern RNA world
Naturally occurring ribozymes include: 1. Co. TC ribozyme 8. Leadzyme 9. Mammalian CPEB 3 ribozyme 2. GIR 1 branching ribozyme 10. r. RNA 3. glm. S ribozyme 11. RNase P 4. Group I and Group II introns 12. Spliceosome 5. Hairpin ribozyme 13. Twister ribozyme 6. Hammerhead ribozyme 14. VS ribozyme 7. HDV ribozyme
Riboswitch
JUNK DNA
Functional structure of human genome (Racaniello, 2014)
• The vast majority (80. 4%) of the human genome participates in at least one biochemical RNA- and/or chromatin-associated event in at least one cell type. ~60% of the genome is involved in transcription.
Many promoters(and enhancers) are bidirectiona Antisense cryptic unstable transcript (CUT)
RNA interference (RNAi) m. RNA Rd. RP ds. RNA Dicer si. RNA, 19 -21 bp RISC
Micro RNAs (mi. RNAs)
mi. RNAs in the regulation of gene expression mi. RNA gene target gene RNAP II pri-mi. RNA pre-mi. RNA RISC Dicer mi. RNA RISC
mi. RNAs are integrated in the regulatory networks Elementary motifs of the network Regulatory network in Drosopila cells Transcription factor gene mi. RNA gene Target gene
Regulatory network in human cells
What is tm. RNA? m. RNA-like and t. RNA-like properties 1. All sequenced eubacteria 2. Some rare mitochondrial and plastidial genomes in simple eukaryotes such as diatoms 3. Some phage genomes
Trans-translation
~1 in 8 expressed genes produced detectable levels of circ. RNA (Jeck et al. 2013)
mi. RNAs Circular RNAs: mi. RNA sponges mi. RNA targets Circ. RNA 5' 3' Splicing
Circ. RNA pecularities - Usually composed of 1 -5 exons - often expressed tissue/developmental stage specific - are predominantly found in the cytoplasm - include long introns and exons - number of circ. RNA can be ten times greater than linear transcript - found in Archea, where they are generated in nonsplicing mechanism - CDR 1 as/Ci. RS-7 targets mi. R-7. It has over 60 mi. R-7 binding sites - some are flanked by inverted repeats
e. RNAs (50 -2000 nucleotides) is produced from bi- or unidirectional transcription of enhancers. Nearly 25% enhancers are transcribed (murine neuron)
Model of e. RNAs effect on gene expression
e. RNA is important for p 53 -dependent transcription
I think you were waiting for this…
Vault organelle is found by cell biologist Nancy Kedersha and biochemist Leonard Rome of the UCLA School of Medicine in 1986 It is identified in mammals, amphibians, avians, molluscs, cnidarian, flatworms.
Weight 13 MDa Structure Vault proteins (95%) Vault RNAs (v. RNAs, also known as vt. RNAs) of 86– 141 bases (5%) – 16 copies of the same RNA
Large intergenic non-coding RNAs (linc. RNAs)
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RNA_world.ppt