The high peak electric fields provided by single-cycle light pulses can be harnessed to manipulate and control charge motion in solid-state systems, resulting in electron emission out of metals and semiconductors [1][2][3][4][5][6] or high harmonics generation in dielectrics 7,8 . These processes are of a non-perturbative character and require precise reproducibility of the electric-field profile 9-14 . Here, we vary the carrier-envelope phase of 6-fs-long near-infrared pulses with pJ-level energy to control electronic transport in a laterally confined nanoantenna with an 8 nm gap. Peak current densities of 50 MA cm -2 are achieved, corresponding to the transfer of individual electrons in a half-cycle period of 2 fs. The observed behaviours are made possible by the strong distortion of the effective tunnelling barrier due to the extreme electric fields that the nanostructure provides and sustains under sub-cycle optical biasing. Operating at room temperature and in a standard atmosphere, the performed experiments demonstrate a robust class of nanoelectronic switches gated by phase-locked optical transients of minute energy content.Present-day high-frequency devices operate in the microwave range, but direct control of electron flow by the electric-field profile of few-cycle optical pulses has recently been demonstrated 12,15 . These experiments were based on strong perturbation of the electronic system in a dielectric material, resulting in a large incoherent background current. Nanoscale confinement of the region biased by the optical transient might avoid a significant influence of nonlinear conductivity in an insulating substrate and result in purely coherent tunnelling currents 16 that may be controlled by the precise shape of the electric field cycles. Our approach to reaching this goal is illustrated in the upper part of Fig. 1a. A single-cycle near-infrared pulse (left) is focused to a nanoscale junction of an electronic circuit (centre) with nonlinear and antisymmetric current-voltage (I-V) characteristics (right). An effective bias then arises when the exciting electric field is cosine-shaped (blue transient and blue dot in the I-V scheme) because its extremum occurs only for one polarity. Consequently, the symmetry of electronic transport is broken, even when integrating over the entire transient, and a net tunnelling current of electrons results through the potential barrier represented by the free-space gap (Fig. 1b,c). Our experiment favours tunnelling over other phenomena such as multiphoton ionization 4 by operating with ultrashort pulses and extreme nanoconfinement of the electric field. No background current exists when the control field is sineshaped (red transient and dot in the I-V characteristics, respectively), because positive and negative polarities now have precisely opposite effects. Consequently, the total current amplitude and direction may be set by varying the carrier-envelope phase (CEP), Δφ CEP , of the pulse. Owing to the single-cycle pulse duration, nanoscale constriction and fi...

Poly(ADP-ribosyl)ation regulates numerous cellular processes like genome maintenance and cell death, thus providing protective functions but also contributing to several pathological conditions. Poly(ADP-ribose) (PAR) molecules exhibit a remarkable heterogeneity in chain lengths and branching frequencies, but the biological significance of this is basically unknown. To unravel structure-specific functions of PAR, we used PARP1 mutants producing PAR of different qualities, i.e. short and hypobranched (PARP1\G972R), short and moderately hyperbranched (PARP1\Y986S), or strongly hyperbranched PAR (PARP1\Y986H). By reconstituting HeLa PARP1 knockout cells, we demonstrate that PARP1\G972R negatively affects cellular endpoints, such as viability, cell cycle progression and genotoxic stress resistance. In contrast, PARP1\Y986S elicits only mild effects, suggesting that PAR branching compensates for short polymer length. Interestingly, PARP1\Y986H exhibits moderate beneficial effects on cell physiology. Furthermore, different PARP1 mutants have distinct effects on molecular processes, such as gene expression and protein localization dynamics of PARP1 itself, and of its downstream factor XRCC1. Finally, the biological relevance of PAR branching is emphasized by the fact that branching frequencies vary considerably during different phases of the DNA damage-induced PARylation reaction and between different mouse tissues. Taken together, this study reveals that PAR branching and chain length essentially affect cellular functions, which further supports the notion of a ‘PAR code’.

Genotoxic stress activates PARP1, resulting in the post-translational modification of proteins with poly(ADP-ribose) (PAR). We genetically deleted PARP1 in one of the most widely used human cell systems, i.e. HeLa cells, via TALEN-mediated gene targeting. After comprehensive characterization of these cells during genotoxic stress, we analyzed structure–function relationships of PARP1 by reconstituting PARP1 KO cells with a series of PARP1 variants. Firstly, we verified that the PARP1\E988K mutant exhibits mono-ADP-ribosylation activity and we demonstrate that the PARP1\L713F mutant is constitutively active in cells. Secondly, both mutants exhibit distinct recruitment kinetics to sites of laser-induced DNA damage, which can potentially be attributed to non-covalent PARP1–PAR interaction via several PAR binding motifs. Thirdly, both mutants had distinct functional consequences in cellular patho-physiology, i.e. PARP1\L713F expression triggered apoptosis, whereas PARP1\E988K reconstitution caused a DNA-damage-induced G2 arrest. Importantly, both effects could be rescued by PARP inhibitor treatment, indicating distinct cellular consequences of constitutive PARylation and mono(ADP-ribosyl)ation. Finally, we demonstrate that the cancer-associated PARP1 SNP variant (V762A) as well as a newly identified inherited PARP1 mutation (F304L\V762A) present in a patient with pediatric colorectal carcinoma exhibit altered biochemical and cellular properties, thereby potentially supporting human carcinogenesis. Together, we establish a novel cellular model for PARylation research, by revealing strong structure–function relationships of natural and artificial PARP1 variants.

Poly(ADP ribos)ylation (PARylation) is an impor tant posttranslational protein modification, and is involved in major cellular processes such as gene regulation and DNA repair. Its dysregulation has been linked to several diseases, including cancer. Despite its importance, methods to observe PARylation dynamics within cells are rare. By following a chemical biology approach, we developed a fluorescent NAD + analogue that proved to be a competitive building block for protein PARylation in vitro and in cells. This allowed us to directly monitor the turnover of PAR in living cells at DNA damage sites after near infrared (NIR) microirradiation. Addi tionally, covalent and noncovalent interactions of selected target proteins with PAR chains were visualized in cells by using FLIM FRET microscopy. Our results open up new opportunities for the study of protein PARylation in real time and in live cells, and will thus contribute to a better under standing of its significance in a cellular context.

We report compact, low-cost and efficient Cr:Colquiriite lasers that are pumped by high brightness tapered laser diodes. The tapered laser diodes provided 1 to 1.2 W of output power around 675 nm, at an electrical-to-optical conversion efficiency of about 30%. Using a single tapered diode laser as the pump source, we have demonstrated output powers of 500 mW and 410 mW together with slope efficiencies of 47% and 41% from continuous wave (cw) Cr:LiSAF and Cr:LiCAF lasers, respectively. In cw mode-locked operation, sub-100-fs pulse trains with average power between 200 mW and 250 mW were obtained at repetition rates around 100 MHz. Upon pumping the Cr:Colquiriite lasers with two tapered laser diodes (one from each side of the crystal), we have observed scaling of cw powers to 850 mW in Cr:LiSAF and to 650 mW in Cr:LiCAF. From the double side pumped Cr:LiCAF laser, we have also obtained ~220 fs long pulses with 5.4 nJ of pulse energy at 77 MHz repetition rate. These are the highest energy levels reported from Cr:Colquiriite so far at these repetition rates. Our findings indicate that tapered diodes in the red spectral region are likely to become the standard pump source for Cr:Colquiriite lasers in the near future. Moreover, the simplified pumping scheme might facilitate efficient commercialization of Cr:Colquiriite systems, bearing the potential to significantly boost applications of cw and femtosecond lasers in this spectral region (750-1000 nm).