Deep generative design of RNA family sequences.

RNA engineering has immense potential to drive innovation in biotechnology and medicine. Despite its importance, a versatile platform for the automated design of functional RNA is still lacking. Here, we propose RNA family sequence generator (RfamGen), a deep generative model that designs RNA family sequences in a data-efficient manner by explicitly incorporating alignment and consensus secondary structure information. RfamGen can generate novel and functional RNA family sequences by sampling points from a semantically rich and continuous representation. We have experimentally demonstrated the versatility of RfamGen using diverse RNA families. Furthermore, we confirmed the high success rate of RfamGen in designing functional ribozymes through a quantitative massively parallel assay. Notably, RfamGen successfully generates artificial sequences with higher activity than natural sequences. Overall, RfamGen significantly improves our ability to design functional RNA and opens up new potential for generative RNA engineering in synthetic biology.

Programmable mammalian translational modulators by CRISPR-associated proteins.

Translational modulation based on RNA-binding proteins can be used to construct artificial gene circuits, but RNA-binding proteins capable of regulating translation efficiently and orthogonally remain scarce. Here we report CARTRIDGE (Cas-Responsive Translational Regulation Integratable into Diverse Gene control) to repurpose Cas proteins as translational modulators in mammalian cells. We demonstrate that a set of Cas proteins efficiently and orthogonally repress or activate the translation of designed mRNAs that contain a Cas-binding RNA motif in the 5'-UTR. By linking multiple Cas-mediated translational modulators, we designed and built artificial circuits like logic gates, cascades, and half-subtractor circuits. Moreover, we show that various CRISPR-related technologies like anti-CRISPR and split-Cas9 platforms could be similarly repurposed to control translation. Coupling Cas-mediated translational and transcriptional regulation enhanced the complexity of synthetic circuits built by only introducing a few additional elements. Collectively, CARTRIDGE has enormous potential as a versatile molecular toolkit for mammalian synthetic biology.

Representation learning applications in biological sequence analysis.

Although remarkable advances have been reported in high-throughput sequencing, the ability to aptly analyze a substantial amount of rapidly generated biological (DNA/RNA/protein) sequencing data remains a critical hurdle. To tackle this issue, the application of natural language processing (NLP) to biological sequence analysis has received increased attention. In this method, biological sequences are regarded as sentences while the single nucleic acids/amino acids or k-mers in these sequences represent the words. Embedding is an essential step in NLP, which performs the conversion of these words into vectors. Specifically, representation learning is an approach used for this transformation process, which can be applied to biological sequences. Vectorized biological sequences can then be applied for function and structure estimation, or as input for other probabilistic models. Considering the importance and growing trend for the application of representation learning to biological research, in the present study, we have reviewed the existing knowledge in representation learning for biological sequence analysis.

Endosomal dysfunction in iPSC-derived neural cells from Parkinson's disease patients with VPS35 D620N.

Mutations in the Vacuolar protein sorting 35 (VPS35) gene have been linked to familial Parkinson's disease (PD), PARK17. VPS35 is a key component of the retromer complex, which plays a central role in endosomal trafficking. However, whether and how VPS35 deficiency or mutation contributes to PD pathogenesis remain unclear. Here, we analyzed human induced pluripotent stem cell (iPSC)-derived neurons from PD patients with the VPS35 D620N mutation and addressed relevant disease mechanisms. In the disease group, dopaminergic (DA) neurons underwent extensive apoptotic cell death. The movement of Rab5a- or Rab7a-positive endosomes was slower, and the endosome fission and fusion frequencies were lower in the PD group than in the healthy control group. Interestingly, vesicles positive for cation-independent mannose 6-phosphate receptor transported by retromers were abnormally localized in glial cells derived from patient iPSCs. Furthermore, we found α-synuclein accumulation in TH positive DA neurons. Our results demonstrate the induction of cell death, endosomal dysfunction and α -synuclein accumulation in neural cells of the PD group. PARK17 patient-derived iPSCs provide an excellent experimental tool for understanding the pathophysiology underlying PD.