**3.1 DNA loops/ bulges and slipped DNA**

DNA loops and bulges are similar non-B DNA structures sharing common features of unpaired bases of different number (Fig. 1). They can be formed in anywhere by any DNA sequence in natural genome, therefore they may be the most frequent non-B DNA conformations in genomes. For example, (CA· TG)n DNA sequences are found to exist everywhere in eukaryotic genomes as of 60 base pairs tracts. (CA· TG)n forms both classical right-handed DNA double helix, and diverse alternative conformations including small DNA loops or bulges (Kladde et al., 1994; Ho, 1994).

Genomic instabilities can also be caused by DNA loops and bulges, which are often seen as slippage instabilities or insertion/deletion ( I/ D) instabilities (Pan, 2004). Proteins that bind DNA loops and bulges are also found and mainly known to be mismatch repair proteins (Parker & Marinus, 1992; Carraway & Marinus, 1993; Fang et al., 2003; Kaliyaperumal et al., 2011).

Fig. 1. DNA Loops and bulges

### **3.2 Branched structures**

A branched DNA structure refers to a non-B DNA secondary structure with structured or unstructured "branch". For example, DNA intermediates appeared in homologous recombination as 3- and 4- way junctures are such branched DNA structures with differently oriented double helix arms. Similarly, flapped DNA structures appeared in processing Okazaki fragment in the lagging strand DNA replication also belong to branched DNA. Branched DNA migrates more slowly than their B-DNA conformation having same molecular weight and base composition. Importantly, branched DNA structures can also make genomic instability when in processing.

#### **3.3 Hairpin/ cruciform and genetic instability**

A hairpin can be formed at one strand of an inverted repeat, whereas a cruciform consists of two hairpin structures, both in each strand at the same position of the DNA (Fig.2)(Courey, 1999). Similarly some tandem arranged trinucleotide repeats such as CAG, CTG, CCG, CGG,

DNA loops and bulges are similar non-B DNA structures sharing common features of unpaired bases of different number (Fig. 1). They can be formed in anywhere by any DNA sequence in natural genome, therefore they may be the most frequent non-B DNA conformations in genomes. For example, (CA· TG)n DNA sequences are found to exist everywhere in eukaryotic genomes as of 60 base pairs tracts. (CA· TG)n forms both classical right-handed DNA double helix, and diverse alternative conformations including small

Genomic instabilities can also be caused by DNA loops and bulges, which are often seen as slippage instabilities or insertion/deletion ( I/ D) instabilities (Pan, 2004). Proteins that bind DNA loops and bulges are also found and mainly known to be mismatch repair proteins (Parker & Marinus, 1992; Carraway & Marinus, 1993; Fang et al., 2003;

A branched DNA structure refers to a non-B DNA secondary structure with structured or unstructured "branch". For example, DNA intermediates appeared in homologous recombination as 3- and 4- way junctures are such branched DNA structures with differently oriented double helix arms. Similarly, flapped DNA structures appeared in processing Okazaki fragment in the lagging strand DNA replication also belong to branched DNA. Branched DNA migrates more slowly than their B-DNA conformation having same molecular weight and base composition. Importantly, branched DNA structures can also

A hairpin can be formed at one strand of an inverted repeat, whereas a cruciform consists of two hairpin structures, both in each strand at the same position of the DNA (Fig.2)(Courey, 1999). Similarly some tandem arranged trinucleotide repeats such as CAG, CTG, CCG, CGG,

**3. The non-B DNA structures and non-B DNA structure-induced genetic** 

**instability** 

**3.1 DNA loops/ bulges and slipped DNA** 

Kaliyaperumal et al., 2011).

Fig. 1. DNA Loops and bulges

make genomic instability when in processing.

**3.3 Hairpin/ cruciform and genetic instability** 

**3.2 Branched structures** 

DNA loops or bulges (Kladde et al., 1994; Ho, 1994).

AAT, ATT etc. can also adopt hairpin structures with mismatched base pairs in the stem (McMurray, 1999; Trotta, et al., 2000).

To form a hairpin/cruciform, DNA duplex needs to be unwound in replication, transcription, and/or DNA repair processing; affording single-stranded repeat sequences the opportunity to base pair with itself in an intramolecular fashion. The term of "cruciform" originates from forming two duplex arms, which adopts either an "open" form, allowing strand migration or a"stacked" (locked) form, where the helices stack on each other (Courey, 1999; Khuu et al., 2006; Lilley, 2010). In both cases, the overall conformation and the intraduplex angles behave like the Holliday junction recombination intermediates (Fig.2A) (Courey, 1999; Khuu et al., 2006;; Lilley, 2010).

#### Fig. 2. Hairpin/cruciform of DNA

Both inverted repeats and tandem arranged trinucleotide repeats were found to be mutagenic, causing genomic instability. Inverted repeats were initially found to cause deletions in *E. coli* (Sinden et al., 1991), and then were seen in humans as (8; 22) (q24.13; q11.21), and many types of t (11; 22) translocations. The breakpoints of these translocation mutations were localized at the center of AT-rich palindromic sequences on 11q23 and 22q11, respectively. So far, t (11; 22) is the only known recurrent, non-Robertsonian translocation in humans, in some cases leads to male infertility and recurrent abortion (Kurahashi et al., 2000, 2006, 2010; Kurahashi & Emanuel, 2001). Furthermore, deletions stimulated by a poly (R.Y) sequence from intron 21 of the polycystic kidney disease 1 gene (PKD1) have also been characterized ( Bacolla et al., 2001;Patel et al., 2004). And a long (CCTG-CAGG)n repeat in *E.coli* was also found to form cruciform (Pluciennik et al., 2002; Dere & Wells, 2006). Interestingly, cruciform-forming inverted repeats have mediated many of the microinversions in evolution that distinguish the human and chimpanzee genomes (Kolb et al., 2009).

In cells, DNA double strand breaks can be derived from cruciform, because hairpin/ cruciform are substrates for several structure-specific nucleases and/ or repair enzymes, such as SbcCD in *E.coli* and Mre11-Rad50 in eukaryotes. The actions of such enzymes make strand breaks, which may result in rearrangements or translocation of chromosomes (Smith, 2008).

In addition, proteins working in nucleotide excisonal repair (NER) can also recognize the helical distortions in hairpin, therefore NER may recognize DNA hairpin to resolve the hairpin in the DNA.

The Gratuitous Repair on Undamaged DNA Misfold 407

may be stabilized by negative supercoiling, modified with phosphorothioate groups, or polyvalent cations such as spermine and spermidine. For the R\* R: Y intramolecular triplexes and T: A\* T and C+: G\* C triplets for the Y\* R: Y intramolecular triplexes are included since

Fig. 3. H-DNA (Star/ Dot marks Hoogsteen hydrogen bonded bases; colon/ line shows

In general, formation of a triplex DNA was a role of sequence, topology (supercoil density), ionic conditions, protein binding, methylation, carcinogen binding, and other factors. Global negative supercoil density acts in concert with local transient waves of topological changes produced by replication or transcription, and both have a critical influence on forming and stabilizing triplex DNA *in vivo*. It has been reported that a higher negative supercoiling destabilized long CTG· CAG, CCG· CGG, and GAA· TTC repeats in *Escherichia coli.* Similarly a 2.5-kb poly (R· Y) tract from the human PKD1 gene lowered the viability of the

Several types of DNA damages induced by H-DNA have been reported, including single and/ or double strand breaks. For example, the endogenous H-DNA forming sequences from the human c-*myc* promoter was shown to be intrinsically mutagenic in mammalian cells because of the generation of either single or double strand breaks in the H-DNA, or near the H-DNA locus. Besides, the single-stranded area, or the triplex region is also a target of various nucleases, resulting in single or DSBs formation, and the increased mutagenesis

Although triplex (H-DNA) DNA occurs mainly at poly (purine·pyrimidine) ((R·Y) n) tracts, it can also be induced to form with the sequence specific DNA recognition and binding of some synthetic triplex-forming oligonucleotides (TFOs) (Casey & Glazer, 2001; Mukherjee & Vasquez, 2011). TFOs bind to the major groove of homopurine-homopyrimidine stretches of double-stranded DNA to induce forming the triplex (Casey & Glazer, 2001; Mukherjee & Vasquez, 2011). During which the duplex DNA may have to undergo helical distortions on TFO binding and the distortions trigger endogenous recombination and repair mechanisms

Indeed it has been reported that formation of TFO-induced triplex can induce sequencespecific DNA damages both in cells and in animals (Chan, et al., 1999; Kalish et al., 2005).

these are considered the most stable triplet combinations.

Watson–Crick hydrogen bonded bases)

host cells (Bacolla et al., 2001; Patel et al., 2004).

or recombination (Wang & Vasquez, 2006).

in the cell (Raghavan et al., 2004, 2005).

Besides, some other proteins were also found to bind the structural elements in cruciforms. For example, HMG proteins, replication initiation protein RepC, cruciform binding protein CBP, and four-way junction resolvases have all been indentified to bind cruciforms (Pearson et al., 1996; Jin et al., 1997; Novac et al., 2002; Lange et al., 2009; lilley, 2010).
