The MPK6-LTF1L1 module regulates lignin biosynthesis in rice through a distinct mechanism from Populus LTF1.

Lignin is a complex polymer that provides structural support and defense to plants. It is synthesized in the secondary cell walls of specialized cells. Through regulates its stability, LTF1 acts as a switch to control lignin biosynthesis in Populus, a dicot plant. However, how lignin biosynthesis is regulated in rice, a monocot plant, remains unclear. By employing genetic, cellular, and chemical approaches, we discovered that LTF1L1, a rice homolog of LTF1, regulates lignin biosynthesis through a distinct mechanism from Populus LTF1. Knockout of LTF1L1 increased lignin synthesis in the sclerenchyma cells of rice stems, while overexpression of LTF1L1 decreased it. LTF1L1 is phosphorylated by OsMPK6 at Ser169, which did not affect its stability but impaired its ability to repress the expression of lignin biosynthesis genes. This was supported by the non-phosphorylated mutant of LTF1L1 (LTF1L1S169A), which displayed a stronger repressive effect on lignin biosynthesis in both rice and Populus. Our findings reveal that LTF1L1 acts as a negative regulator of lignin biosynthesis via a distinct mechanism from that of LTF1 in Populus and highlight the evolutionary diversity in the regulation of lignin biosynthesis in plants.

Screening genome-editing knockouts reveals the receptor-like kinase ASX role in regulations of secondary xylem development in Populus.

In trees, secondary xylem development is essential for the growth of perennial stem increments. Many signals regulate the process of development, but our knowledge of the molecular components involved in signal transduction is still limited. In this study, we identified Attenuation of Secondary Xylem (ASX) knockouts by screening genome-editing knockouts of xylem-expressed receptor-like kinases (RLKs) in Populus. The ASX role in secondary xylem development in Populus was discovered using biochemical, cellular, and genomic analyses. The ASX knockout plants had abnormal secondary stem growth but had little effect on shoot apical primary growth. ASX and SOMATIC EMBRYOGENESIS RECEPTOR KINASE (SERK)2/4 were co-precipitated in developing xylem. Through their interaction, ASX is phosphorylated by SERK. Transcriptome analysis of developing xylem revealed that ASX deficiency inhibited the transcriptional activity of genes involved in xylem differentiation and secondary cell wall formation. By forming a complex, ASX and SERK may function as a signaling module for signal transduction required in the regulation of secondary xylem development in trees. This study shows that ASX, which encodes a RLKs, is required for secondary xylem development and sheds light on regulatory signals found in tree stem secondary growth.

Two MADS-box genes regulate vascular cambium activity and secondary growth by modulating auxin homeostasis in Populus.

In trees, stem secondary growth depends on vascular cambium proliferation activity and subsequent cell differentiation, in which an auxin concentration gradient across the cambium area plays a crucial role in regulating the process. However, the underlying molecular mechanism for the establishment of auxin concentration is not fully understood. In this study, we identified two function-unknown MADS-box genes, VCM1 and VCM2, which are expressed specifically in the vascular cambium and modulate the subcellular homeostasis of auxin. Simultaneous knockdown of both VCM1 and VCM2 enhanced vascular cambium proliferation activity and subsequent xylem differentiation. Overexpression of VCM1 suppressed vascular cambium activity and wood formation by regulating PIN5 expression, which tuned the soluble auxin concentration in the vascular cambium area. This study reveals the role of VCM1 and VCM2 in regulating the proliferation activity of the vascular cambium and secondary growth by modulating the subcellular auxin homeostasis in Populus.

Cell-Specific Suppression of 4-Coumarate-CoA Ligase Gene Reveals Differential Effect of Lignin on Cell Physiological Function in Populus.

Lignin is a main component of the secondary cell wall in vessels and fibers of xylem tissue. However, the significance of lignin in cell physiology during plant growth is unclear. In this study, we generated lignin-modified Populus via cell-specific downregulation of the 4-coumarate-CoA ligase gene (4CL). The transgenic plants with selective lignin modification in vessel elements or fiber cells allowed us to investigate how lignin affects the physiology of vessel or fiber cells in relation to plant growth. Results showed that vessel-specific suppression of lignin biosynthesis resulted in deformed vessels and normal fibers, while fiber-specific suppression of lignin biosynthesis led to less-lignified fibers and normal vessels. Further analyses revealed that the efficiency of long distance water transport was severely affected in transgenics with vessel-specific lignin modification, while minimal effect was detected in transgenics with fiber-specific lignin modification. Vessel-specific lignin reduction led to high susceptibility to drought stress and poor growth in field, likely due to vessel defects in long distance transport of water. The distinct physiological significance of lignin in different cell types provides insights into the selective modification of lignin for improvement of lignocellulosic biomass utilization.

High-flow nasal cannula oxygen therapy versus non-invasive ventilation for chronic obstructive pulmonary disease patients after extubation: a multicenter, randomized controlled trial.

BACKGROUND: High-flow nasal cannula (HFNC) oxygen therapy is being increasingly used to prevent post-extubation hypoxemic respiratory failure and reintubation. However, evidence to support the use of HFNC in chronic obstructive pulmonary disease (COPD) patients with hypercapnic respiratory failure after extubation is limited. This study was conducted to test if HFNC is non-inferior to non-invasive ventilation (NIV) in preventing post-extubation treatment failure in COPD patients previously intubated for hypercapnic respiratory failure. METHODS: COPD patients with hypercapnic respiratory failure who were already receiving invasive ventilation were randomized to HFNC or NIV at extubation at two large tertiary academic teaching hospitals. The primary endpoint was treatment failure, defined as either resumption of invasive ventilation or switching to the other study treatment modality (NIV for patients in the NFNC group or vice versa). RESULTS: Ninety-six patients were randomly assigned to the HFNC group or NIV group. After secondary exclusion, 44 patients in the HFNC group and 42 patients in the NIV group were included in the analysis. The treatment failure rate in the HFNC group was 22.7% and 28.6% in the NIV group-risk difference of - 5.8% (95% CI, - 23.8-12.4%, p = 0.535), which was significantly lower than the non-inferior margin of 9%. Analysis of the causes of treatment failure showed that treatment intolerance in the HFNC group was significantly lower than that in the NIV group, with a risk difference of - 50.0% (95% CI, - 74.6 to - 12.9%, p = 0.015). One hour after extubation, the mean respiratory rates of both groups were faster than their baseline levels before extubation (p < 0.050). Twenty-four hours after extubation, the respiratory rate of the HFNC group had returned to baseline, but the NIV group was still higher than the baseline. Forty-eight hours after extubation, the respiratory rates of both groups were not significantly different from the baseline. The average number of daily airway care interventions in the NIV group was 7 (5-9.3), which was significantly higher than 6 (4-7) times in the HFNC group (p = 0.006). The comfort score and incidence of nasal and facial skin breakdown of the HFNC group was also significantly better than that of the NIV group [7 (6-8) vs 5 (4-7), P < 0.001] and [0 vs 9.6%, p = 0.027], respectively. CONCLUSION: Among COPD patients with severe hypercapnic respiratory failure who received invasive ventilation, the use of HFNC after extubation did not result in increased rates of treatment failure compared with NIV. HFNC also had better tolerance and comfort than NIV. TRIAL REGISTRATION: chictr.org ( ChiCTR1800018530 ). Registered on 22 September 2018, http://www.chictr.org.cn/usercenter.aspx.

Superfine grinding induced amorphization and increased solubility of α-chitin.

As the most common structure of chitin, α-chitin is insoluble in common aquatic and organic solvents, and is very difficult to be processed due to its highly ordered crystalline structure and the large number of intermolecular and intramolecular hydrogen bonds. Amorphization of α-chitin has been proved to be a valid measure for improving its subsequent functionalization efficiency and depolymerization yield. In this study, superfine grinding (SFG) was introduced to make α-chitin amorphous, and it was found that SFG effectively reduced the particle size, changed the microstructure, and significantly reduced the crystallinity of α-chitin. Chitin with crystallinity as low as 8.39 % was obtained after 60 min of SFG treatment, and the amorphous chitin became readily dissolved in 10 % NaOH solution after one round of freezing-thawing process. As continuous manner could be employed, SFG might be a powerful and efficient method for preparing amorphous chitin to help its processing and modification of various applications.

Fibre-specific regulation of lignin biosynthesis improves biomass quality in Populus.

Lignin is a major component of cell wall biomass and decisively affects biomass utilisation. Engineering of lignin biosynthesis is extensively studied, while lignin modification often causes growth defects. We developed a strategy for cell-type-specific modification of lignin to achieve improvements in cell wall property without growth penalty. We targeted a lignin-related transcription factor, LTF1, for modification of lignin biosynthesis. LTF1 can be engineered to a nonphosphorylation form which is introduced into Populus under the control of either a vessel-specific or fibre-specific promoter. The transgenics with lignin suppression in vessels showed severe dwarfism and thin-walled vessels, while the transgenics with lignin suppression in fibres displayed vigorous growth with normal vessels under phytotron, glasshouse and field conditions. In-depth lignin structural analyses revealed that such cell-type-specific downregulation of lignin biosynthesis led to the alteration of overall lignin composition in xylem tissues reflecting the population of distinctive lignin polymers produced in vessel and fibre cells. This study demonstrates that fibre-specific suppression of lignin biosynthesis resulted in the improvement of wood biomass quality and saccharification efficiency and presents an effective strategy to precisely regulate lignin biosynthesis with desired growth performance.

Duration of cardiac arrest requires different ventilation volumes during cardiopulmonary resuscitation in a pig model.

There are few studies examining the ventilation strategies recommended by current CPR guidelines. We investigated the influence of different minute volume applying to untreated cardiac arrest with different duration, on resuscitation effects in a pig model. 32 Landrace pigs with 4 or 8 min (16 pigs each) ventricular fibrillation (VF) randomly received two ventilation strategies during CPR. "Guideline" groups received mechanical ventilation with a tidal volume of 7 ml/kg and a frequency of 10/min, while "Baseline" groups received a tidal volume (10 ml/kg) and a frequency used at baseline to maintain an end-tidal PCO2 (PETCO2) between 35 and 40 mmHg before VF. Mean airway pressures and intrathoracic pressures (PIT) in the Baseline-4 min group were significantly higher than those in the Guideline-4 min group (all P < 0.05). Similar results were observed in the 8 min pigs, except for no significant difference in minimal PIT and PETCO2 during 10 min of CPR. Venous pH and venous oxygen saturation were significantly higher in the Baseline-8 min group compared to the Guideline-8 min group (all P < 0.05). Aortic pressure in the Baseline-8 min group was higher than in the Guideline-8 min group. Seven pigs in each subgroup of 4 min VF models achieved the return of spontaneous circulation (ROSC). Higher ROSC was observed in the Baseline-8 min group than in the Guideline-8 min group (87.5% vs. 37.5%, P = 0.039). For 4 min VF but not 8 min VF, a guideline-recommended ventilation strategy had satisfactory results during CPR. A higher minute ventilation resulted in better outcomes for subjects with 8 min of untreated VF through thoracic pump.

A xylem-produced peptide PtrCLE20 inhibits vascular cambium activity in Populus.

In trees, lateral growth of the stem occurs through cell divisions in the vascular cambium. Vascular cambium activity is regulated by endogenous developmental programmes and environmental cues. However, the underlying mechanisms that regulate cambium activity are largely unknown. Genomic, biochemical and genetic approaches were used here to elucidate the role of PtrCLE20, a CLAVATA3 (CLV3)/embryo surrounding region (ESR)-related peptide gene, in the regulation of lateral growth in Populus. Fifty-two peptides encoded by CLE genes were identified in the genome of Populus trichocarpa. Among them PtrCLE20 transcripts were detected in developing xylem while the PtrCLE20 peptide was mainly localized in vascular cambium cells. PtrCLE20 acted in repressing vascular cambium activity indicated by that upregulation of PtrCLE20 resulted in fewer layers of vascular cambium cells with repressed expression of the genes related to cell dividing activity. PtrCLE20 peptide also showed a repression effect on the root growth of Populus and Arabidopsis, likely through inhibiting meristematic cell dividing activity. Together, the results suggest that PtrCLE20 peptide, produced from developing xylem cells, plays a role in regulating lateral growth by repression of cambium activity in trees.

High flow nasal cannula oxygen therapy versus non-invasive ventilation for chronic obstructive pulmonary disease with acute-moderate hypercapnic respiratory failure: an observational cohort study.

Background: High-flow nasal cannula (HFNC) oxygen therapy in acute hypoxic respiratory failure is becoming increasingly popular. However, evidence to support the use of HFNC in acute respiratory failure (ARF) with hypercapnia is limited. Methods: Chronic obstructive pulmonary disease (COPD) patients with moderate hypercapnic ARF (arterial blood gas pH 7.25-7.35, PaCO2>50 mmHg) who received HFNC or non-invasive ventilation (NIV) in the intensive care uint from April 2016 to March 2018 were analyzed retrospectively. The endpoint was treatment failure, defined as either invasive ventilation, or a switch to the other study treatment (NIV for patients in the NFNC group, and vice-versa), and 28-day mortality. Results: Eighty-two COPD patients (39 in the HFNC group and 43 in the NIV group) were enrolled in this study. The mean age was 71.8±8.2 and 54 patients (65.9%) were male. The treatment failed in 11 out of 39 patients with HFNC (28.2%) and in 17 of 43 patients with NIV (39.5%) (P=0.268). No significant differences were found for 28-day mortality (15.4% in the HFNC group and 14% in the NIV group, P=0.824). During the first 24 hrs of treatment, the number of nursing airway care interventions in the HFNC group was significantly less than in the NIV group, while the duration of device application was significantly longer in the HFNC group (all P<0.05). Skin breakdown was significantly more common in the NIV group (20.9% vs 5.1%, P<0.05). Conclusion: Among COPD patients with moderate hypercarbic ARF, the use of HFNC compared with NIV did not result in increased rates of treatment failure, while there were fewer nursing interventions and skin breakdown episodes reported in the HFNC group.

[Benefits and risks of stress ulcer prevention with proton pump inhibitors for critical patients: an observational cohort study with 1 972 patients].

OBJECTIVE: To investigate the benefits and risks of stress ulcer prevention (SUP) using proton pump inhibitors (PPI) for critical patients. METHODS: The clinical data of adult critically ill patients admitted to the intensive care unit (ICU) of Northern Jiangsu People's Hospital from January 2016 to December 2018 were retrospectively analyzed. All patients who were treated with PPI for SUP within the first 48 hours after ICU admission were enrolled in the SUP group. Those who not received PPI were enrolled in the control group. A one-to-one propensity score matching (PSM) was performed to control for potential biases. The gender, age, underlying diseases, main diagnosis of ICU, drug use before ICU admission, sequential organ failure score (SOFA) at ICU admission, risk factors of stress ulcer (SU) and PPI usage were recorded. The end point was the incidence of gastrointestinal bleeding, hospital acquired pneumonia, Clostridium difficile infection and 30-day mortality. Kaplan-Meier survival curves were plotted, and survival analysis was performed using the log-rank test. RESULTS: 1 972 critical patients (788 in the SUP group and 1 184 in the control group) were enrolled, and each group enrolled 358 patients after PSM. Prior to PSM, compared with the control group, the SUP group had older patients, more underlying diseases, higher proportion of acute coronary syndrome (ACS), acute cerebrovascular disease, acute exacerbation of chronic obstructive pulmonary disease (AECOPD) and poisoning in main diagnosis of ICU, more serious illness, and more risk factors of SU, indicating that ICU physicians were more likely to prescribe SUP for these patients. The incidence of gastrointestinal bleeding in the SUP group was significantly lower than that in the control group [1.8% (14/788) vs. 3.7% (44/1 184), P < 0.05], while the incidence of hospital acquired pneumonia and 30-day mortality were significantly higher than those in the control group [6.6% (52/788) vs. 3.5% (42/1 184), 17.9% (141/788) vs. 13.1% (155/1 184), both P < 0.01]. There was no significant difference in the incidence of Clostridium difficile infection between the SUP group and the control group [2.9% (23/788) vs. 1.8% (21/1 184), P > 0.05]. After the propensity scores for age, underlying diseases, severity of illness and SU risk factors were matched, there was no significant difference in the incidence of gastrointestinal bleeding or 30-day mortality between the SUP group and the control group [2.2% (8/358) vs. 3.4% (12/358), 15.9% (57/358) vs. 13.7% (49/358), both P > 0.05], but the incidence of hospital acquired pneumonia in the SUP group was still significantly higher than that in the control group [6.7% (24/358) vs. 3.1% (11/358), P < 0.05]. Kaplan-Meier survival curve analysis showed that the 30-day cumulative survival rate of the SUP group was significantly lower than that of the control group before the PSM (log-rank test: χ2 = 9.224, P = 0.002). There was no significant difference in the 30-day cumulative survival rate between the two groups after PSM (log-rank test: χ2 = 0.773, P = 0.379). CONCLUSIONS: For critical patients, the use of PPI for SUP could not significantly reduce the incidence of gastrointestinal bleeding and mortality, but increase the risk of hospital acquired pneumonia.

Phosphorylation of LTF1, an MYB Transcription Factor in Populus, Acts as a Sensory Switch Regulating Lignin Biosynthesis in Wood Cells.

Lignin is specifically deposited in plant secondary cell walls, and initiation of lignin biosynthesis is regulated by a variety of developmental and environmental signals. However, the mechanisms governing the regulation of lignin biosynthesis remain to be elucidated. In this study, we identified a lignin biosynthesis-associated transcription factor (LTF) from Populus, LTF1, which binds the promoter of a key lignin biosynthetic gene encoding 4-coumarate-CoA ligase (4CL). We showed that LTF1 in its unphosphorylated state functions as a regulator restraining lignin biosynthesis. When LTF1 becomes phosphorylated by PdMPK6 in response to external stimuli such as wounding, it undergoes degradation through a proteasome pathway, resulting in activation of lignification. Expression of a phosphorylation-null mutant version of LTF1 led to stable protein accumulation and persistent attenuation of lignification in wood cells. Taken together, our study reveals a mechanism whereby LTF1 phosphorylation acts as a sensory switch to regulate lignin biosynthesis in response to environmental stimuli. The discovery of novel modulators and mechanisms modifying lignin biosynthesis has important implications for improving the utilization of cell-wall biomass.

A HD-ZIP III gene, PtrHB4, is required for interfascicular cambium development in Populus.

Wood production is dependent on the activity of the vascular cambium, which develops from the fascicular and interfascicular cambia. However, little is known about the mechanisms controlling how the vascular cambium is developed in woody species. Here, we show that PtrHB4, belonging to the Populus HD-ZIP III family, plays a critical role in the process of vascular cambium development. PtrHB4 was specifically expressed in shoot tip and stem vascular tissue at an early developmental stage. Repression of PtrHB4 caused defects in the development of the secondary vascular system due to failures in interfascicular cambium formation. By contrast, overexpression of PtrHB4 induced cambium activity and xylem differentiation during secondary vascular development. Transcriptional analysis of PtrHB4 repressed plants indicated that auxin response and cell proliferation were affected in the formation of the interfascicular cambium. Taken together, these results suggest that PtrHB4 is required for interfascicular cambium formation to develop the vascular cambium in woody species.

Formation of wood secondary cell wall may involve two type cellulose synthase complexes in Populus.

Cellulose biosynthesis is mediated by cellulose synthases (CesAs), which constitute into rosette-like cellulose synthase complexe (CSC) on the plasma membrane. Two types of CSCs in Arabidopsis are believed to be involved in cellulose synthesis in the primary cell wall and secondary cell walls, respectively. In this work, we found that the two type CSCs participated cellulose biosynthesis in differentiating xylem cells undergoing secondary cell wall thickening in Populus. During the cell wall thickening process, expression of one type CSC genes increased while expression of the other type CSC genes decreased. Suppression of different type CSC genes both affected the wall-thickening and disrupted the multilaminar structure of the secondary cell walls. When CesA7A was suppressed, crystalline cellulose content was reduced, which, however, showed an increase when CesA3D was suppressed. The CesA suppression also affected cellulose digestibility of the wood cell walls. The results suggest that two type CSCs are involved in coordinating the cellulose biosynthesis in formation of the multilaminar structure in Populus wood secondary cell walls.

Suppression of PtrDUF579-3 Expression Causes Structural Changes of the Glucuronoxylan in Populus.

DUF579 (domain unknown function 579) genes have been reported to play diverse roles in cell wall biosynthesis, such as in glucuronoxylan (GX) synthesis. As GX is a major type of hemicelluloses in hard wood species, how DUF579 genes function in wood formation remains to be demonstrated in planta. This study reports a Populus DUF579 gene, PtrDUF579-3, which is characterized for its function in wood cell wall formation. PtrDUF579-3 is localized in Golgi apparatus and catalyzes methylation of the glucuronic acid (GlcA) in GX biosynthesis. Suppression of PtrDUF579-3 expression in Populus caused a reduction in both the GlcA side chain number and GlcA side chain methylation on the GX backbone. The modified GX polymer through PtrDUF579-3 suppression was more susceptible to acid treatment and the PtrDUF579-3 suppressed plants displayed enhanced cellulose digestibility. These results suggest that PtrDUF579-3 is involved in GX biosynthesis and GX structure can be modified through PtrDUF579-3 suppression in Populus.

Accelerated partial breast irradiation with intensity-modulated radiotherapy is feasible for chinese breast cancer patients.

PURPOSE: Several accelerated partial breast irradiation (APBI) techniques are being investigated in patients with early-stage breast cancer. The present study evaluated the feasibility, early toxicity, initial efficacy, and cosmetic outcomes of accelerated partial breast intensity-modulated radiotherapy (IMRT) for Chinese female patients with early-stage breast cancer after breast-conserving surgery. METHODS: A total of 38 patients met the inclusion criteria and an accelerated partial breast intensity-modulated radiotherapy (APBI-IMRT) plan was designed for each patient. The prescription dose was 34 Gy in 10 fractions, 3.4 Gy per fraction, twice a day, in intervals of more than 6 hours. RESULTS: Of the 38 patients, six patients did not meet the planning criteria. The remaining 32 patients received APBI-IMRT with a mean target volume conformity index of 0.67 and a dose homogeneity index of 1.06. The median follow-up time was 53 months and no local recurrence or distant metastasis was detected. The most common acute toxicities observed within 3 months after radiotherapy were erythema, breast edema, pigmentation, and pain in the irradiated location, among which 43.8%, 12.5%, 31.3%, and 28.1% were grade 1 toxicities, respectively. The most common late toxicities occurring after 3 months until the end of the follow-up period were breast edema, pigmentation, pain in the irradiated location, and subcutaneous fibrosis, among which 6.2%, 28.1%, 21.9%, and 37.5% were grade 1 toxicities, respectively. Thirty-one patients (96.8%) had fine or excellent cosmetic outcomes, and only one patient had a poor cosmetic outcome. CONCLUSION: It is feasible for Chinese females to receive APBI-IMRT after breast conserving surgery. The radiotherapeutic toxicity is acceptable, and both the initial efficacy and cosmetic outcomes are good.

Diverse roles of PtrDUF579 proteins in Populus and PtrDUF579-1 function in vascular cambium proliferation during secondary growth.

DUF579 (domain of unknown function 579) family proteins contain a DUF579 domain structure but vary greatly in their overall sequence similarity. Several DUF579 proteins have been found to play a role in cell wall biosynthesis in Arabidopsis, while DUF579 family genes have not yet been systematically investigated in Populus. In this study, the Populus DUF579 family proteins were found to be localized in different cell types and subcellular locations. The diverse expression patterns of the proteins indicate that they may perform different functions in Populus. Among the DUF579 family members, PtrDUF579-1 is found to be specifically expressed in vascular cambium zone cells where it is localized in the Golgi apparatus. Suppression of PtrDUF579-1 expression reduced plant height and stem diameter size. Cambium cell division and xylem tissue growth was inhibited while secondary cell wall formation was unchanged in PtrDUF579-1 suppressed plants. Cell walls analysis showed that the composition of the pectin fraction of the cambium cell wall was altered while other polysaccharides were not affected in PtrDUF579-1 suppressed plants. This observation suggest cambium expressed PtrDUF579-1 may affect cell wall biosynthesis and be involved in cambium cell proliferation in Populus. Overall, DUF579 family proteins play a diverse set of roles in Populus.

PtrKOR1 is required for secondary cell wall cellulose biosynthesis in Populus.

KORRIGAN (KOR), encoding an endo-1,4-ß-glucanase, plays a critical role in the cellulose synthesis of plant cell wall formation. KOR sequence orthologs are duplicated in the Populus genome relative to Arabidopsis. This study reports an expression analysis of the KOR genes in Populus. The five PtrKOR genes displayed different expression patterns, suggesting that they play roles in different developmental processes. Through RNAi suppression, results demonstrated that PtrKOR1 is required for secondary cell wall cellulose formation in Populus. Together, the results suggest that the PtrKOR genes may play distinct roles in association with cell wall formation in different tissues.

Intron-mediated alternative splicing of WOOD-ASSOCIATED NAC TRANSCRIPTION FACTOR1B regulates cell wall thickening during fiber development in Populus species.

Alternative splicing is an important mechanism involved in regulating the development of multicellular organisms. Although many genes in plants undergo alternative splicing, little is understood of its significance in regulating plant growth and development. In this study, alternative splicing of black cottonwood (Populus trichocarpa) wood-associated NAC domain transcription factor (PtrWNDs), PtrWND1B, is shown to occur exclusively in secondary xylem fiber cells. PtrWND1B is expressed with a normal short-transcript PtrWND1B-s as well as its alternative long-transcript PtrWND1B-l. The intron 2 structure of the PtrWND1B gene was identified as a critical sequence that causes PtrWND1B alternative splicing. Suppression of PtrWND1B expression specifically inhibited fiber cell wall thickening. The two PtrWND1B isoforms play antagonistic roles in regulating cell wall thickening during fiber cell differentiation in Populus spp. PtrWND1B-s overexpression enhanced fiber cell wall thickening, while overexpression of PtrWND1B-l repressed fiber cell wall thickening. Alternative splicing may enable more specific regulation of processes such as fiber cell wall thickening during wood formation.

PtrCel9A6, an endo-1,4-ß-glucanase, is required for cell wall formation during xylem differentiation in populus.

Endo-1,4-ß-glucanases (EGases) are involved in many aspects of plant growth. Our previous study found that an EGase, PtrCel9A6, is specifically expressed in differentiating xylem cells during Populus secondary growth. In this study, the xylem-specific PtrCel9A6 was characterized for its role in xylem differentiation. The EGase is localized on the plasma membrane with catalytic domain toward the outside cell wall, hydrolyzing amorphous cellulose. Suppression of PtrCel9A6 expression caused secondary cell wall defects in xylem cells and significant cellulose reduction in Populus. Heterologous expression of PtrCel9A6 in Arabidopsis enhanced plant growth as well as increased fiber cell length. In addition, introduction of PtrCel9A6 into Arabidopsis resulted in male sterility due to defects in anther dehiscence. Together, these results demonstrate that PtrCel9A6 plays a critical role in remodeling the 1,4-ß-glucan chains in the wall matrix and is required for cell wall thickening during Populus xylem differentiation.