Biology subjects

Dalla Benetta, E.

Publications and source records attributed to Dalla Benetta, E..

5 recordsLinked to original sources

bioRxiv2024

Synthetic type III-E CRISPR-Cas effectors for programmable RNA-targeting

The recent discovery of the type III-E CRISPR-Cas effector class has reshaped our fundamental understanding of CRISPR-Cas evolution and classification. Type III-E effectors are composed of several Cas7-like domains and a single Cas11-like domain naturally fused together to create a single polypeptide capable of programmably targeting and degrading RNA. Here we identify a novel composition of a type III-E-like effector composed of Cas7-like and a Cas1-like domain, that can be engineered into an active chimeric RNA-targeting Cas effector and presents a new function of Cas1 in RNA-targeting. Furthermore, we demonstrate a unique modularity of type III-E effectors by methodically substituting domains between orthogonal type III-E proteins to engineer compact synthetic Cas effectors. We refine our methods to generate several compact effectors for programmable RNA-targeting in mammalian cells. Cas7-S represents a new understanding of type III-E architecture and modularity, and provides a platform for engineering genome engineering technologies from the blueprint of nature.

molecular biology↗

bioRxiv2023

Engineered Antiviral Sensor Targets Infected Mosquitoes

Escalating vector disease burdens pose significant global health risks, so innovative tools for targeting mosquitoes are critical. We engineered an antiviral strategy termed REAPER (vRNA Expression Activates Poisonous Effector Ribonuclease) that leverages the programmable RNA-targeting capabilities of CRISPR Cas13 and its potent collateral activity. Akin to a stealthy Trojan Horse hiding in stealth awaiting the presence of its enemy, REAPER remains concealed within the mosquito until an infectious blood meal is up taken. Upon target viral RNA infection, REAPER activates, triggering programmed destruction of its target arbovirus such as chikungunya. Consequently, Cas13 mediated RNA targeting significantly reduces viral replication and its promiscuous collateral activity can even kill infected mosquitoes. This innovative REAPER technology adds to an arsenal of effective molecular genetic tools to combat mosquito virus transmission. One-Sentence SummaryEngineered Cas13-based antiviral sensor kills infected mosquitoes upon activation from arbovirus infection.

bioengineering↗

bioRxiv2020

Germline mutagenesis of Nasonia vitripennis through ovarian delivery of CRISPR-Cas9 ribonucleoprotein

CRISPR/Cas9 gene editing is a powerful technology to study the genetics of rising model organisms, such as the jewel wasp Nasonia vitripennis. However, current methods involving embryonic microinjection of CRISPR reagents are challenging. Delivery of Cas9 ribonucleoprotein into female ovaries is an alternative that has only been explored in a small handful of insects, such as mosquitoes and whiteflies. Here, we developed a simple protocol for germline gene editing by injecting Cas9 ribonucleoprotein in adult N. vitripennis females using either ReMOT control (Receptor-Mediated Ovary Transduction of Cargo) or BAPC (Branched Amphiphilic Peptide Capsules) as ovary delivery methods. We demonstrate efficient delivery of protein cargo such as EGFP and Cas9 into developing oocytes via P2C peptide and BAPC. Additionally, somatic and germline gene editing have been demonstrated. This approach will greatly facilitate CRISPR-applied genetic manipulation in this and other rising model organisms.

molecular biology↗

bioRxiv2019

Genome Elimination Mediated by Gene Expression from a Selfish Chromosome

Numerous plants and animals harbor selfish B chromosomes that \"drive,\" or transmit themselves at higher-than-Mendelian frequencies, despite long-term fitness costs to the organism. Currently it is unknown how B chromosome drive is mediated, and whether B-gene expression plays a role. We used modern sequencing technologies to analyze the fine-scale sequence composition and expression of Paternal Sex Ratio (PSR), a B chromosome in the jewel wasp Nasonia vitripennis. PSR causes female-to-male conversion by destroying the sperms hereditary material in young embryos in order to drive. Using RNA interference, we show that testis-specific expression of a PSR-linked gene, named haploidizer, facilitates this genome elimination-and-sex conversion effect. Haploidizer shares homology with a gene in Candidatus cardinium, a bacterial symbiont that also induces genome elimination in its insect host.\n\nOne Sentence Summaryhaploidizer mediates B chromosome drive

evolutionary biology↗

bioRxiv2019

Latitudinal variation in circadian rhythmicity in Nasonia vitripennis

Many physiological processes of living organisms show circadian rhythms, governed by an endogenous clock. This clock has a genetic basis and is entrained by external cues such as light and temperature. Other physiological processes exhibit seasonal rhythms, that are also responsive to light and temperature. We previously reported a natural latitudinal cline of photoperiodic diapause induction in the parasitic wasp Nasonia vitripennis in Europe and a correlated haplotype frequency for the circadian clock gene period (per). To evaluate if this correlation is reflected in circadian behaviour, we investigated circadian locomotor activity of seven populations from the cline. We found that the proportion of rhythmic males is higher than females in constant darkness, and that mating decreased rhythmicity of both sexes. Only for virgin females, the free running period ({tau}) increased weakly with latitude. Wasps from the most southern locality had an overall shorter free running rhythm and earlier onset, peak and offset of activity during the 24 h period, than wasps from the northernmost locality. We evaluate this variation in rhythmicity as a function of period haplotype frequencies in the populations and discuss its functional significance in the context of local adaptation.

evolutionary biology↗