This study presents single-nucleus multi-omic profiling of human placental syncytiotrophoblasts (STB) to identify cellular trajectories during pregnancy. Using integrated single-nucleus RNA sequencing (snRNA-seq) and single-nucleus ATAC sequencing (snATAC-seq), researchers analyzed 101,543 placental nuclei from early and late pregnancy. The findings reveal dynamic heterogeneity and developmental trajectories of STB nuclei, their correspondence with human trophoblast stem cell (hTSC)-derived STB, and the transcription factors associated with diverse STB nuclear lineages. The study identifies master transcription factors (TFs) such as MITF, STAT5A, CEBPB, and FOSL2, and their gene regulatory networks. The results highlight the heterogeneity of human STB and provide a valuable resource for interpreting pregnancy complications. The placenta, a critical organ in pregnancy, supports fetal development by supplying nutrients and oxygen and removing waste. It develops during implantation when trophoblasts differentiate from the trophectoderm of the blastocyst. The syncytiotrophoblast (STB), a multinucleated cell type, plays a vital role in placental function. The study shows that STB nuclei exhibit dynamic heterogeneity and developmental trajectories during pregnancy, with distinct subtypes identified in early and late pregnancy. These subtypes are associated with different functions, including hormone synthesis, immunological defense, and active transport. The study also reveals that STB nuclei in early pregnancy have distinct gene expression and chromatin accessibility patterns, with subclusters showing different stages of maturation. The analysis identified two mature STB subtypes, eSTB mature 1 and eSTB mature 2, with distinct gene expression profiles. The study further demonstrates that hTSC-derived STB and trophoblast organoids can mimic the heterogeneity and regulatory mechanisms of STB in early pregnancy. In late pregnancy, the study reveals that STB nuclei exhibit different subtypes, with STB mature 1 dominating. The analysis shows that STB nuclei in late pregnancy have distinct gene expression and chromatin accessibility patterns, with subclusters showing different stages of maturation. The study also identifies key TFs involved in STB development and function, such as STAT6, STAT5A, and MITF, and their roles in regulating gene expression and function. Overall, the study provides a comprehensive analysis of STB nuclear heterogeneity during pregnancy, highlighting the dynamic developmental trajectories and regulatory mechanisms of STB. The findings offer insights into the role of STB in maintaining a successful pregnancy and provide a valuable resource for understanding pregnancy complications.This study presents single-nucleus multi-omic profiling of human placental syncytiotrophoblasts (STB) to identify cellular trajectories during pregnancy. Using integrated single-nucleus RNA sequencing (snRNA-seq) and single-nucleus ATAC sequencing (snATAC-seq), researchers analyzed 101,543 placental nuclei from early and late pregnancy. The findings reveal dynamic heterogeneity and developmental trajectories of STB nuclei, their correspondence with human trophoblast stem cell (hTSC)-derived STB, and the transcription factors associated with diverse STB nuclear lineages. The study identifies master transcription factors (TFs) such as MITF, STAT5A, CEBPB, and FOSL2, and their gene regulatory networks. The results highlight the heterogeneity of human STB and provide a valuable resource for interpreting pregnancy complications. The placenta, a critical organ in pregnancy, supports fetal development by supplying nutrients and oxygen and removing waste. It develops during implantation when trophoblasts differentiate from the trophectoderm of the blastocyst. The syncytiotrophoblast (STB), a multinucleated cell type, plays a vital role in placental function. The study shows that STB nuclei exhibit dynamic heterogeneity and developmental trajectories during pregnancy, with distinct subtypes identified in early and late pregnancy. These subtypes are associated with different functions, including hormone synthesis, immunological defense, and active transport. The study also reveals that STB nuclei in early pregnancy have distinct gene expression and chromatin accessibility patterns, with subclusters showing different stages of maturation. The analysis identified two mature STB subtypes, eSTB mature 1 and eSTB mature 2, with distinct gene expression profiles. The study further demonstrates that hTSC-derived STB and trophoblast organoids can mimic the heterogeneity and regulatory mechanisms of STB in early pregnancy. In late pregnancy, the study reveals that STB nuclei exhibit different subtypes, with STB mature 1 dominating. The analysis shows that STB nuclei in late pregnancy have distinct gene expression and chromatin accessibility patterns, with subclusters showing different stages of maturation. The study also identifies key TFs involved in STB development and function, such as STAT6, STAT5A, and MITF, and their roles in regulating gene expression and function. Overall, the study provides a comprehensive analysis of STB nuclear heterogeneity during pregnancy, highlighting the dynamic developmental trajectories and regulatory mechanisms of STB. The findings offer insights into the role of STB in maintaining a successful pregnancy and provide a valuable resource for understanding pregnancy complications.