Injectable recombinant collagen hydrogels (RCHs) are crucial in biomedical applications. Culture conditions play an important role in the preparation of hydrogels. However, determining the characteristics of hydrogels under certain conditions and determining the optimal conditions swiftly still remain challenging tasks. In this study, a machine learning approach was introduced to explore the correlation between hydrogel characteristics and culture conditions and determine the optimal culture conditions. The study focused on four key factors as independent variables: initial substrate concentration, reaction temperature, pH level, and reaction time, while the dependent variable was the elastic modulus of the hydrogels. To analyze the impact of these factors on the elastic modulus, four mathematical models were employed, including multiple linear regression (ML), decision tree (DT), support vector machine (SVM), and neural network (NN). The theoretical outputs of NN were closest to the actual values. Therefore, NN proved to be the most suitable model. Subsequently, the optimal culture conditions were identified as a substrate concentration of 15% (W/V), a reaction temperature of 4 °C, a pH of 7.0, and a reaction time of 12 h. The hydrogels prepared under these specific conditions exhibited a predicted elastic modulus of 15,340 Pa, approaching that of natural elastic cartilage.

Organogenesis in plants occurs across all stages of the life cycle. Although previous studies have identified many genes as important for either vegetative or reproductive development at the RNA level, global information on translational and post-translational levels remains limited. In this study, six Arabidopsis stages/organs were analyzed using quantitative proteomics and phosphoproteomics, identifying 2187 non-redundant proteins and evidence for 1194 phosphoproteins. Compared to the expression observed in cauline leaves, the expression of 1445, 1644, and 1377 proteins showed greater than 1.5-fold alterations in stage 1–9 flowers, stage 10–12 flowers, and open flowers, respectively. Among these, 294 phosphoproteins with 472 phosphorylation sites were newly uncovered, including 275 phosphoproteins showing differential expression patterns, providing molecular markers and possible candidates for functional studies. Proteins encoded by genes preferentially expressed in anther (15), meiocyte (4), or pollen (15) were enriched in reproductive organs, and mutants of two anther-preferentially expressed proteins, acos5 and mee48, showed obviously reduced male fertility with abnormally organized pollen exine. In addition, more phosphorylated proteins were identified in reproductive stages (1149) than in the vegetative organs (995). The floral organ-preferential phosphorylation of GRP17, CDC2/CDKA.1, and ATSK11 was confirmed with western blot analysis. Moreover, phosphorylation levels of CDPK6 and MAPK6 and their interacting proteins were elevated in reproductive tissues. Overall, our study yielded extensive data on protein expression and phosphorylation at six stages/organs and provides an important resource for future studies investigating the regulatory mechanisms governing plant development.

Pollen wall formation requires precise coordination between tapetum differentiation, metabolism, and programmed cell death. In Arabidopsis, the microspore-derived peptide CLE19 restricts tapetal activity to maintain pollen wall homeostasis, yet how CLE19 signaling achieves developmental specificity and robustness remains unclear. Here, we identify the receptor-like kinase BARELY ANY MERISTEM 3 (BAM3) as an additional receptor for CLE19. Genetic, cytological, biochemical, and transcriptomic analyses showed that disruption of BAM3 signaling impairs tapetum differentiation, secretory homeostasis, and pollen exine patterning, resulting in selective transcriptional reprogramming during anther development. Comparative transcriptomic analyses reveal that BAM3 mediates a distinct subset of CLE19-responsive genes, including both AMS-dependent pathways governing tapetal degeneration and exine biosynthesis, and AMS-independent programs associated with flavonoid metabolism and pollen wall development. Biochemical assays, structure-guided mutagenesis, and AlphaFold3 modeling further support CLE19-dependent assembly of BAM3-SERK1/2 receptor complex, revealing a conserved molecular framework for CLE19 perception that is distinct from, yet complementary to, the previously characterized CLE19–PXL1–SERK1/2 receptor module. Together, these findings establish a dual-receptor architecture for CLE19 signaling in which BAM3 mediates a transcriptionally distinct branch of the CLE19 pathway. More broadly, this work demonstrates how combinatorial peptide-receptor usage expands the signaling capacity of a single developmental peptide to coordinate robust male reproductive development.

Background: RNA helicases participate in multiple aspects of RNA metabolism, but the functions of most helicases in malaria parasites remain poorly understood. PfDDX1 is a conserved DEAD-box RNA helicase whose role during the asexual blood-stage development of Plasmodium falciparum has not been defined. Methods: Endogenous PfDDX1 fusion-protein lines were generated using CRISPR/Cas9-mediated genome editing. PfDDX1 expression and subcellular distribution were examined by Western blotting and immunofluorescence assays. A PfDDX1-HA-ddFKBP conditional knockdown line was used to assess parasite proliferation, intraerythrocytic developmental cycle (IDC) progression, merozoite production, and erythrocyte invasion. Comparative RNA sequencing was performed using morphologically stage-matched ring- and schizont-stage parasites. RNA immunoprecipitation sequencing (RIP-seq) was used to identify PfDDX1-associated RNAs. Results: PfDDX1 contains conserved helicase-associated domains and showed a predominantly nuclear or perinuclear distribution during asexual blood-stage development. PfDDX1 was detected at the ring, trophozoite, and schizont stages, with the highest protein abundance in trophozoites. Conditional PfDDX1 knockdown was associated with earlier progression through the IDC and increased merozoite production per schizont beginning in the first cycle. Parasitemia increased cumulatively over successive cycles and became significantly higher in the fourth cycle, whereas merozoite invasion efficiency was unchanged. RNA-seq revealed stage-dependent changes in transcript abundance, with relatively limited changes at the ring stage and a broader upregulated response at the schizont stage. Schizont-stage upregulated genes were enriched for functions associated with merozoite development and invasion-related cellular structures, including rhoptries, micronemes, the inner membrane complex, and the cell surface. RIP-seq identified stage-associated PfDDX1-enriched RNAs, and integration with RNA-seq revealed a subset of PfDDX1-associated transcripts whose abundance increased following knockdown. Conclusions: PfDDX1 knockdown is associated with altered intraerythrocytic development, increased merozoite production, and stage-dependent changes in transcript abundance in P. falciparum . Although several invasion-associated transcripts were upregulated, erythrocyte invasion efficiency was not detectably altered. These findings support a role for PfDDX1 in RNA-associated regulation during asexual blood-stage development, while the precise transcriptional or post-transcriptional mechanisms require further investigation.