The study evaluated 550 winter wheat cultivars and advanced lines from China’s Northern and Huang-Huai winter wheat zones (Supplementary Table S1), where freezing damage frequently occurs.

Field trials were conducted over three consecutive growing seasons (2022–2023, 2023–2024, and 2024–2025) in Gaocheng, Hebei Province (114.76° E, 37.95° N). The experiment was arranged in a randomized complete block design (RCBD) with two blocks. Within each block, all materials were randomly assigned to experimental plots. Each plot consisted of one 3-m-long rows with 25 cm spacing between rows and 2 cm spacing between plants within a row.

During the 2024–2025 season, plant architecture (PA) was assessed in early December, when seedling growth ceased and mean daily temperature was near 0 °C. This timing corresponds to the maximum tillering stage, where tiller development is complete, ensuring consistent classification. The tiller angle, a key component of PA, was measured as the angle between the outermost fully expanded leaf of tiller and an imaginary vertical axis (Figure 1A) using a crop angle meter (Zhejiang Top Cloud-agriculture Technology Co., Ltd., Hangzhou, Zhejiang Province). Five randomly selected individuals were dug up one by one from each row and placed on the device platform for photography. Image recognition was then conducted, and subsequently, an angle value was generated (Figure 1B). The mean PA value from two replicates was used for final analysis. Based on these angle measurements, seedling PA was categorized into five distinct types according to the International Union for the Protection of New Varieties of Plants (UPOV, 2022) classification standards(https://www.upov.int/documents/d/upov/tg-documents-en-tg003.pdf) (Supplementary Table S2). Representative phenotypes for the erect and prostrate germplasm are shown in Figure 2.

Following winter dormancy, freeze tolerance (FT) was evaluated during the green-up period in mid-March, when mean daily temperatures reached approximately 6 °C. Meteorological data obtained from the weather station of the Institute of Cereal and Oil Crops, Hebei Academy of Agricultural and Forestry Sciences, which is situated within 2 km of the experimental plots(Supplementary Table S3). Fluctuations in ambient air temperature and soil temperature at a depth of 5 cm during the seedling stage across the three growing seasons are depicted in Figures 3, 4, respectively. FT was assessed using two established metrics: severity of leaf necrosis (SLN) and mortality rate of shoots (MRS). The SLN method involves visual evaluation of leaf necrosis during early spring green-up when new heart leaves reach 1–2 cm in length according to Sun et al. (2013). The extent of above-ground necrosis was recorded and used to classify FT into five levels (Supplementary Table S4). For MRS, a 0.5 m section was excavated from each row. The total numbers of plants, shoots, and dead shoots were counted according to Liao et al. (2007). MRS was calculated for each replicate as the percentage of deceased shoots out of the total number of shoots. The mean MRS value from two replicates was used for final analysis and classified into five levels (Supplementary Table S5).

Data management and basic descriptive statistics, including mean values, standard deviations, and percentage for PA, SLN, and MRS, were performed using Microsoft Excel 2019. All subsequent statistical analyses were conducted in SPSS (version 29.0, IBM Corp.). Pearson correlation analysis was used to examine relationships among PA, SLN, and MRS. A one-way analysis of variance (ANOVA) was employed to test for significant differences in SLN and MRS across the three growing seasons, with post-hoc comparisons conducted using Tukey’s HSD test where appropriate. Statistical significance was defined at α = 0.05, and all tests were two-tailed. Effect sizes are reported where applicable to complement significance testing.

PA was evaluated for 550 wheat accessions during the 2024–2025 growing season. The PA distribution across the population approximated a normal distribution (Figure 5). Among the five PA grades assessed, Grade 5 was most prevalent, comprising 30.18% (n = 166) of the accessions. The remaining grades accounted for 13.45% (Grade 1), 20.19% (Grade 3), 21.45% (Grade 7), and 14.73% (Grade 9), respectively.

The correlation between seedling PA and FT varied substantially across years. In both 2022–2023 and 2023-2024, no significant correlation was detected between PA and SLN. The lack of relationship in 2022–2023 coincided with a sudden temperature drop prior to seedling cold acclimation, while in 2023-2024, protective snow cover likely mitigated freezing damage(Supplementary Tables S3–1, S3-2). In the 2024–2025 season, an extremely significant negative correlation emerged between PA and SLN (P < 0.01; Table 2), indicating that more prostrate growth habits were associated with reduced leaf necrosis. The variations across different years indicate that winter temperatures are not the sole predominant factor affecting the PA-SLN relationship. MRS showed no significant correlation with PA across all three growing seasons, suggesting that ultimate shoot survival remains independent of seedling architecture.

Over the three growing seasons, several freeze-tolerant varieties were identified across the diverse panel of 550 accessions (as detailed in Supplementary Table S1). Notably, these tolerant varieties were present across all PA categories, including erect (e.g., Zhongmai 7158), semi-erect (e.g., Nongda 1980), intermediate (e.g., Jimai 479), semi-prostrate (e.g., Xingmai 29), and prostrate (e.g., Henong 405) types. This finding indicates that genetic freezing tolerance can be achieved independently of specific PA, highlighting the complexity of this trait and the likelihood of multiple, compensatory physiological and molecular mechanisms underlying survival. Consequently, these results imply that breeding for FT should not be restricted to selecting a single, idealized PA. Instead, efforts should focus on integrating tolerance genes within a range of suitable, locally adapted plant architectures.

The seedling PA of the 550 winter wheat cultivars and advanced lines was predominantly characterized as intermediate to semi-prostrate. This growth habit appears to balance ecological adaptation with photosynthetic efficiency. The lower canopy profile reduces exposure to chilling winds, while maintaining adequate leaf area for light capture. Such an architectural balance likely supports both winter survival and biomass accumulation during early growth stages. Research in durum wheat has identified prostrate juvenile growth as a valuable trait for enhancing weed competitiveness and water conservation through improved ground coverage (Marone et al., 2020). Wheat leaves retained a complete photosynthetic apparatus, induced sustained nonphotochemical quenching (NPQ) during the cold period in winter, and relaxed NPQ rapidly during the warm period. The photosynthetic apparatus of wheat switched quickly between photosynthetic carbon assimilation and the photoprotective state during the winter (Li et al., 2025). Thus, selection for intermediate seedling PA represents a promising strategy for developing winter wheat varieties with enhanced FT and climate resilience without compromising yield potential.

A significant negative correlation between SLN and PA grade emerged during the 2024–2025 growing season, revealing that more prostrate seedling sustained less leaf necrosis. This finding aligns with observations in durum wheat, where prostrate growth habits reduced frost exposure (Marone et al., 2020). The genetic regulation of prostrate growth involves loci linked to vernalization and frost resistance genes on chromosome 5A (Roberts, 1990), resulting in compact plants characterized by increased tillering, narrower leaves, and reduced elongation (Li et al., 2016). Evidence shows that there are differences in the accumulation of cold-resistant substances between erect and prostrate wheat seedlings. Transcriptomics and nontargeted metabolomics analyses revealed that lignin biosynthesis, ABC transporters, sucrose and glycerophospholipid metabolism were key pathways underlying the differentiation of erect and prostrate growth habits in the wheat overwintering period (Cheng et al., 2025).

The morphological adaptation of a prostrate growth habit is frequently part of an evolutionary suite of traits in cold-adapted plants, working together to reduce physical exposure, protect meristems, and support the physiological and biochemical processes involved in cold acclimation.

The magnitudes of FT among the 550 germplasms varied significantly across the three growing seasons, demonstrating the strong influence of climatic conditions on winter survival. Global climate change cause longer and warmer autumns, thus negatively affecting the quality of cold acclimation and reducing the FT of winter wheat (Rapacz et al., 2017). The development of FT depends critically on successful cold acclimation, a process increasingly disrupted by climate change as prolonged autumn warmth delays hardening, ultimately reducing cold hardiness. This effect was evident in late November 2022, when a sharp temperature drop occurred before acclimation was complete, resulting in significantly higher SLN and MRS values compared to subsequent seasons. Recent research corroborates that extended exposure to mild autumn temperatures, while potentially boosting shoot biomass, can concurrently diminish freezing resistance (Vaitkevičiūtė et al., 2022).

Snow cover also substantially moderated frost injury, as demonstrated in mid-December 2023 when continuous snowfall insulated the soil and minimized freeze-thaw cycling. The protective role of snow exhibits spatial dependence, with even minor variations in snow depth causing substantial differences in survival rates within single fields (Hayhoe and Andrews, 1999). For instance, winter wheat survival at the Motala site in Sweden improved significantly with snow depth increases from 0 to 5 cm (Bergjord-Olsen et al., 2018). These results collectively highlight that both temperature regimes during acclimation and snow cover dynamics are key determinants of field-based FT expression.

The shoot apical meristem (SAM), situated near the soil surface within unelongated internodes, benefits from soil insulation and accumulate protective carbohydrates. This study confirmed that leaves are more freezing-sensitive than tillers, with intact tillers capable of regenerating new shoots even following complete leaf necrosis. The critical role of the crown region (2–3 cm below soil surface) in frost resistance is well established (Bao et al., 2021), with winter-hardy cultivars like ‘Nongda 211’ achieving LT₅₀ temperatures as low as -21.2 °C (Wu et al., 2024). These results collectively demonstrate how soil microclimate and growth habits interact to enhance overwintering survival by protecting crown meristems.

During the winter season, atmospheric temperatures generally decrease to levels lower than those of the soil, rendering the leaves of the aerial plant parts more vulnerable to freeze damage compared to the subterranean SAM. In this study, MRS assessment demonstrated consistently higher tolerance ratings. SLN is more sensitive to cold weather than MRS, and it can further distinguish the FT of different varieties. Other alternate methods, such as LT50 assays, controlled freeze tests, electrolyte leakage, OJIP, NDVI, could improve discrimination on FT (Mu et al., 2015; Grabelnych et al., 2014; Rapacz et al., 2015; Shammi et al., 2023; Sun et al., 2025). While MRS serves as an unambiguous endpoint for plant survival, its correlation with SLN was inconsistent, suggesting tissue-level damage does not always predict whole-plant fate. Thus, the MRS provides a more reliable evaluation of genetic freezing tolerance compared to other methods.