A kinesin-13 family kinesin in Trypanosoma brucei regulates cytokinesis and cytoskeleton morphogenesis by promoting microtubule bundling.

The early branching eukaryote Trypanosoma brucei divides uni-directionally along the longitudinal cell axis from the cell anterior toward the cell posterior, and the cleavage furrow ingresses along the cell division plane between the new and the old flagella of a dividing bi-flagellated cell. Regulation of cytokinesis in T. brucei involves actomyosin-independent machineries and trypanosome-specific signaling pathways, but the molecular mechanisms underlying cell division plane positioning remain poorly understood. Here we report a kinesin-13 family protein, KIN13-5, that functions downstream of FPRC in the cytokinesis regulatory pathway and determines cell division plane placement. KIN13-5 localizes to multiple cytoskeletal structures, interacts with FPRC, and depends on FPRC for localization to the site of cytokinesis initiation. Knockdown of KIN13-5 causes loss of microtubule bundling at both ends of the cell division plane, leading to mis-placement of the cleavage furrow and unequal cytokinesis, and at the posterior cell tip, causing the formation of a blunt posterior. In vitro biochemical assays demonstrate that KIN13-5 bundles microtubules, providing mechanistic insights into the role of KIN13-5 in cytokinesis and posterior morphogenesis. Altogether, KIN13-5 promotes microtubule bundle formation to ensure cleavage furrow placement and to maintain posterior cytoskeleton morphology in T. brucei.

Study of Anatomical Changes of the Inferior Nasal Passage After Le Fort I Osteotomy With Superior Repositioning.

The relationship between postoperative morphological changes in the inferior nasal cavity and inferior turbinate after Le Fort I osteotomy remains unclear. This study aimed to investigate how the bone volume of the inferior turbinate affects contact with the inferior nasal cavity of patients who underwent superior repositioning. We evaluated the 3-dimensional relationship between the anatomical changes in the inferior nasal passage before and after surgery in 51 patients who underwent Le Fort I osteotomy with an elevation of >4.0 mm in the first molar. The soft tissue and bone volumes of the inferior turbinate and airway volume of the inferior nasal passage were calculated using Proplan CMF 3.0 and compared according to the size of the bone volume of the inferior turbinate. In addition, we reclassified the maxillary movements in the pitch direction and compared the results. The contact rates of the postoperative inferior nasal airway and the inferior turbinate in the large-bone group was 72.3% and that in the small-bone group was 40.0% in the χ2 test. The reduction in the inferior nasal passage volume was significantly greater in the large-bone group (pitch+) than in the small-bone group (pitch+). For patients with well-developed bony tissue of the inferior turbinate, caution is advised if the maxillary elevation is ≥4.0 mm, because the possibility of postoperative obstruction of the inferior nasal passages exist, which may lead to deterioration of nasal ventilation.

Postoperative stability of bioresorbable plates made of 85:15 poly (L-lactide-co-glycolide) in Le Fort I osteotomy.

OBJECTIVE: Recently, RapidSorb plates (DePuy Synthes) made of 85.15 poly (L-lactide-co-glycolide) have been used for orthognathic surgery; however, reports regarding their effectiveness are limited. We aimed to compare the postoperative stability of RapidSorb plates, RapidSorb combined with titanium (MOJ plates), and MOJ plates in patients who underwent Le Fort I osteotomy at Tokyo Medical and Dental University Hospital. STUDY DESIGN: The use of RapidSorb in the maxilla is a load-sharing application and therefore constitutes an approved indication. Discrepancies in the maxillary positions were measured using postoperative computed tomography data at 1 week and 1 year using the centroid method 3-dimensionally. Treatment with RapidSorb alone showed a more vertical discrepancy in the maxilla treatment with MOJ and RapidSorb+MOJ. The RapidSorb4 group was subdivided into 2 groups (under and over 1.0-mm) based on the change in the maxillary centroid. RESULTS: The bone gap at the lateral border of the piriform aperture was significantly larger in the over-1.0-mm group than in the 1.0-mm group. CONCLUSIONS: The fixation of RapidSorb alone is not appropriate in load-bearing and unstable applications but is not contraindicated for load-sharing indications. Fixation with RapidSorb combined with MOJ was clinically effective, with results similar to titanium plate-only fixation regarding postoperative stability.

Risk factors for postoperative facial swelling at 1 week after bimaxillary orthognathic surgery in Class III patients.

OBJECTIVE: To quantify facial swelling at 1 week after Le Fort I osteotomy and bilateral sagittal splitting ramus osteotomy in Class III patients and to identify factors contributing to the swelling based on clinical, morphologic, and surgical variables. STUDY DESIGN: Data from 63 patients were examined in this single-center, retrospective study. Facial swelling was quantitatively measured by superimposing computed tomography data taken in the supine position at 1 week and 1 year postoperatively and extracting the area of maximum intersurface distance. Age, sex, body mass index, thickness of subcutaneous tissue, and of masseter muscle, maxillary length (A-VRP), mandibular length (B-VRP), and posterior maxillary height (U6-HRP), surgical movement (ΔA-VRP, ΔB-VRP, ΔU6-HRP), drainage method, and usage of facial bandages were examined. Multiple regression analysis was performed using the above factors. RESULTS: The median swelling at 1 week postoperatively was 8.35 IQR (5.99-11.47) mm. Multiple regression analysis revealed three factors that were significantly associated with facial swelling: Use of postoperative facial bandages (P=0.03), masseter muscle thickness (P=0.03), and ΔB-VRP (P=0.04). CONCLUSION: Absence of a facial bandage, thin masseter muscle, and large horizontal mandibular movement are risk factors for facial swelling at 1 week postoperatively.

Effects of Perioperative Oral Management in Patients with Cancer.

Perioperative oral management (POM) is used to prevent pneumonia in patients with cancer. However, the factors that expose hospitalized patients to increased risk of developing pneumonia remain unclear. For example, no study to date has compared the incidence of pneumonia in hospitalized patients by cancer primary lesion, or POM implementation, or not. We determined which patients were most likely to benefit from POM and examined the effects of POM on pneumonia prevention and mortality. In a total of 9441 patients with cancer who underwent surgery during hospitalization, there were 8208 patients in the No POM group, and 1233 in the POM group. We examined between-group differences in the incidence of pneumonia and associated outcomes during hospitalization. There was no significant between-group difference in the incidence of pneumonitis, however, patients with lung, or head and neck cancers, demonstrated a lower incidence of postoperative pneumonia. Among patients with lung and pancreatic cancers, mortality was significantly lower in the POM group. POM appears effective at reducing the risk of postoperative pneumonia in patients with certain cancers. Further, mortality was significantly lower in patients with lung and pancreatic cancers who received POM; hence, POM may be an effective adjuvant therapy for patients with cancer.

Risk factors of neurosensory disturbances at 1 year postoperatively after bilateral sagittal split osteotomy.

OBJECTIVE: To investigate morphologic and surgical risk factors causing neurosensory disturbances (NSDs) after bilateral sagittal split osteotomy (BSSO). STUDY DESIGN: A total of 237 patients (with 474 sides) who underwent BSSO were followed up for 1 year. Parameters examined included age, sex, asymmetry, mandibular movement direction, mandible cutting devices, split type, intraoperative exposure of the inferior alveolar nerve (IAN), contact between the IAN and screw, distance between mandibular canal and inner surface of the cortical bone (distance A), distance from lateral osteotomy to mental foramen (distance B), and NSD at 1 year postoperatively. RESULTS: NSD was observed in 62 (13.1%) sides of 51 patients. Exploratory factor analysis determined 4 factors (factor 1: distance A; factor 2: direction of mandibular movement; factor 3: distance B and cutting devices; factor 4: IAN exposure). Logistic regression analysis was performed using the above factors and age, sex, and asymmetry, making a total of 7 variables. Age, factor 1, and factor 4 were significant predictors of NSD. CONCLUSIONS: Advanced age, close distance between mandibular canal and inner surface of the cortical bone, and IAN intraoperative exposure are risk factors for NSD 1 year postoperatively. Cases at high risk for NSD must be treated with great care.

Polo-like kinase and Aurora B kinase phosphorylate and cooperate with the CIF1-CIF2 complex to promote cytokinesis initiation in Trypanosoma brucei.

Cytokinesis in eukaryotes is regulated by a Polo-like kinase-mediated and Aurora B kinase-mediated signalling pathway that promotes the assembly of the actomyosin contractile ring, a cytokinesis machinery conserved across evolution from yeast to humans. Trypanosoma brucei, an early divergent parasitic protozoan, employs an actomyosin-independent mechanism for its unusual cytokinesis that is controlled by a regulatory pathway comprising the Polo-like kinase TbPLK, the Aurora B kinase TbAUK1 and multiple trypanosomatid-specific regulators. However, whether any of these trypanosomatid-specific regulators function as substrates of TbPLK and/or TbAUK1 and how they cooperate with TbPLK and TbAUK1 to promote cytokinesis remain unknown. Here, we demonstrate that TbPLK and TbAUK1 phosphorylate the cytokinesis regulators CIF1 and CIF2 on multiple sites within their intrinsically disordered regions. We further show that TbPLK localization depends on its interaction with CIF1 from S/G2 phases, that TbPLK maintains CIF1 and CIF2 localization from G2 phase until early mitosis, and that TbAUK1 maintains CIF1 and CIF2 localization from late mitosis. Finally, we demonstrate that the cytokinesis regulators CIF4 and FPRC are not substrates of TbPLK and TbAUK1, and that they function upstream of TbPLK and TbAUK1 in the cytokinesis regulatory pathway. Together, these results provide insights into the functional interplay and the order of actions between the two protein kinases and the trypanosomatid-specific cytokinesis regulators in T. brucei.

Study of Morphological Changes in the Inferior Turbinate After Le Fort I Osteotomy.

OBJECTIVE: There are cases in which patients complain of nasal obstruction after Le Fort I osteotomy, but the relationship with postoperative morphological changes in the nasal cavity, including the septum and inferior turbinate, is not clear. STUDY DESIGN: The authors evaluated the three-dimensional relationship of the morphological changes in the inferior turbinate before and after surgery in 84 patients who underwent Le Fort I osteotomy. Three classifications were made according to superior amount of maxillary movement at the base of nasal cavity. RESULTS: The high elevation group (4.0 mm or more) had 31 sides, the moderate elevation group had 93 sides, and the low elevation group (less than 2.0 mm) had 44 sides. The volume of inferior turbinate was 76.9 ± 12.8% of that before surgery in the high elevation group. The high- and moderate-elevation groups had significantly higher changes than the low elevation group, and the rate of contact between inferior turbinate and nasal cavity floor was 67.7%. CONCLUSIONS: After Le Fort I osteotomy, the volume of inferior turbinate tissue decreased in proportion to the amount of elevation of the maxilla. Although the soft tissue volume may be reduced due to adaptation of respiratory function, the inferior nasal passage was not completely ventilated in the high elevation group. If the elevation exceeds 4.0 mm counterclockwise with maxillary movement, it is necessary to consider the concomitant inferior turbinate resection because it may lead to nasal obstruction.

Changes in the condylar volume and skeletal relapse following orthognathic surgery in patients with dentofacial deformity: A retrospective study.

OBJECTIVE: To evaluate the relationship between the changes in condylar volume and maxillofacial skeletal morphology according to sex as well as the relationship between condylar volume reduction and skeletal relapse in patients who underwent orthognathic surgery. METHODS: Ninety-five patients were categorized into skeletal Class III, Class II, and facial asymmetry groups. Computed tomography scans taken preoperatively and at 1 year postoperatively were used for quantitative measurement. RESULTS: Postoperative condylar volume was reduced in both the Class II group and the deviated side of the asymmetry group. Both female and Class II deformity were significant predictors of postoperative reduction in the condylar volume. There was a significant correlation between skeletal relapse and postoperative change in condylar volume in the Class II group. CONCLUSION: Postoperative condylar resorption may be associated with preoperative maxillofacial skeletal morphology and sex and also with skeletal relapse in the Class II group.

Structural Domains of CIF3 Required for Interaction with Cytokinesis Regulatory Proteins and for Cytokinesis Initiation in Trypanosoma brucei.

Cytokinesis in Trypanosoma brucei occurs unidirectionally from the anterior toward the posterior through mechanisms distinct from those of its human host and is controlled by a signaling pathway comprising evolutionarily conserved and trypanosome-specific regulatory proteins. The mechanistic roles and the functional interplay of these cytokinesis regulators remain poorly understood. Here, we investigate the requirement of the structural motifs in the trypanosome-specific cytokinesis regulator CIF3 for the initiation of cytokinesis, the interaction with other cytokinesis regulators, and the recruitment of CIF3-interacting proteins to the cytokinesis initiation site. We demonstrate that the internal and C-terminal coiled-coil motifs, but not the N-terminal coiled-coil motif, of CIF3 play essential roles in cytokinesis and interact with distinct cytokinesis regulators. CIF3 interacts with TbPLK, CIF1, CIF4, and FPRC through the N-terminal and C-terminal coiled-coil motifs and with KAT80 through all three coiled-coil motifs. The C-terminal coiled-coil motif of CIF3 is required for the localization of CIF3 and all of its interacting proteins, and additionally, the internal coiled-coil motif of CIF3 is required for KAT80 localization. Conversely, all the CIF3-interacting proteins are required to maintain CIF3 at the cytokinesis initiation site at different cell cycle stages. These results demonstrate that CIF3 cooperates with multiple interacting partner proteins to promote cytokinesis in T. brucei. IMPORTANCE Cytokinesis is the final stage of cell division and is regulated by a signaling pathway conserved from yeast to humans. Cytokinesis in Trypanosoma brucei, an early-branching protozoan parasite causing human sleeping sickness, is regulated by mechanisms that are distinct from those of its human host, employing a number of trypanosome-specific regulatory proteins to cooperate with evolutionarily conserved regulators. The functional interplay of these cytokinesis regulators is still poorly understood. In this work, we investigated the structural requirement of the trypanosome-specific cytokinesis regulator CIF3 for the initiation of cytokinesis, the interaction with other cytokinesis regulatory proteins, and the recruitment of CIF3-interacting proteins. We demonstrated that different structural motifs of CIF3 played distinct roles in cytokinesis, interacted with distinct cytokinesis regulatory proteins, and were required for the recruitment of distinct cytokinesis regulatory proteins. These findings provided novel insights into the cooperative roles of cytokinesis regulators in promoting cytokinesis in T. brucei.

What does oral care mean to society?

Oral care is defined in a narrow sense as cleaning of the teeth, oral cavity, and dentures, and in a broad sense as the maintenance of oral functions (feeding, swallowing, chewing, speech, aesthetics, etc.), dental treatment, feeding and swallowing training, and articulation training. In the past, it was recognized as simply cleaning the mouth, but the concept of oral care has gradually expanded, and many studies and surveys have been conducted in cooperation with various other professions. As a result, oral health care is involved not only in the prevention of pneumonia, but also in the onset and suppression for severity of diabetes, cardiovascular diseases, some malignant tumors, cerebrovascular diseases, rheumatoid arthritis, dementia, etc. It is also a powerful supportive therapy in cancer treatment. In the terminal stages of life, oral health care can help people to maintain their dignity by continuing to consume food orally until the end of their lives, and in times of disaster, oral health care has been found to be as important as attention to deep vein thrombosis. It has also been found to be effective in preventing severe diseases such as COVID-19. And, although it has not been discussed much, it has been found to have medical and economic benefits such as reducing the duration of hospitalization and treatment costs. This article reviews the results of research to date.

Retrospective analysis of schwannoma in the oral and maxillofacial region: clinicopathological characteristics and specific pathology of ancient change.

Schwannomas are benign tumours that originate from the Schwann cells of the nerve. Despite the frequency of their occurrence in the head and neck, data relating to their clinicopathological features in the region are limited. This study reviews the clinicopathological characteristics and specific pathology of ancient (degenerative) change in 40 cases in the oral and maxillofacial region. Medical records were reviewed of the 40 cases treated at Tokyo Medical and Dental University Hospital Faculty of Dentistry between 2000 and 2020. The most frequently involved site was the tongue, and the average tumour size was 13.2 mm. Degenerative changes were observed in eight cases. All tumours were completely excised through biopsy or local excision, and no recurrence was observed on clinical follow up. Statistical analyses revealed significant associations (p < 0.05) between ancient change and tumour size, and between ancient change and the apparent diffusion coefficient (ADC) value derived from diffusion-weighted magnetic resonance imaging (MRI). Analysis suggests that schwannoma progresses over a long period and subsequently undergoes secondary ancient changes. Pathological events such as cystic formation and haemorrhage that are associated with ancient changes, cause hypocellularity and lead to high ADC values. These values may indicate ancient change and should be considered when distinguishing schwannoma from other lesions.

Short-Term Stability After Segmental Le Fort I Maxillary Impaction Surgery With Mandibular Autorotation in Seven High-Angle Class II Patients: A Case Series.

PURPOSE: To retrospectively evaluate skeletal stability after Le Fort I maxillary impaction surgery and mandibular autorotation without bilateral sagittal split osteotomy (BSSO) in high-angle class II patients. MATERIALS AND METHODS: Seven female high-angle class II patients who underwent maxillary impaction surgery and mandibular autorotation without bilateral sagittal split osteotomy were included in this study. Surgical changes and relapse were measured on lateral cephalograms taken preoperatively and at 1 month, 6 months and 1 year postoperatively. RESULTS: The horizontal movement of the maxilla at point A was 5.8 ±â€Š3.3 mm backward, and the upward movement at the posterior nasal spine was 3.3 ±â€Š1.4 mm. The mean horizontal change at point A during the 1-year follow-up period was 0.1 ±â€Š0.2 mm, and the vertical change at posterior nasal spine was 0.2 ±â€Š1.3 mm, which were not statistically significant. The horizontal surgical change at point B was 4.0 ±â€Š1.8 mm forward and the vertical surgical change at point B was 4.7 ±â€Š1.8 mm upward. Postoperative relapse was 10.9% and 13.7% in the horizontal and vertical directions, respectively. CONCLUSIONS: Le Fort I maxillary impaction surgery with mandibular autorotation may be 1 of the suitable procedures for high-angle class II patients.

Long-term follow-up after arthroplasty for pediatric temporomandibular joint ankylosis performed before the critical period of mandibular growth: A case report.

INTRODUCTION AND IMPORTANCE: Pediatric temporomandibular joint (TMJ) ankylosis can lead to significant difficulties in opening the mouth, as well as stunted mandibular growth resulting in a small mandible and facial asymmetry. For pediatric TMJ ankylosis, the ideal time to perform TMJ mobilization in order to achieve standard mandibular growth is unclear. CASE PRESENTATION: An 11-year-old boy with limited mouth opening was referred to our hospital. The patient had previously sustained a fracture of the left articular process of the mandible. Clinical examination revealed bony ankylosis of the left TMJ, and the condylar head was dislocated anteromedially. The bony ankylosis was removed at the age of 12 years. Mouth-opening exercises were started postoperatively. His mouth opening recovered without the development of severe facial asymmetry, and the dislocated condylar process served as a new joint and provided good jaw function until the most recent follow-up at the age of 21 years. CLINICAL DISCUSSION: It is practical to perform surgical intervention after the child has achieved some growth and at a time when the surgery would not interfere with jaw development because the mouth opening can be improved by postoperative physical therapy. No interpositional implant was used because strict postoperative mouth opening exercises and the displaced condylar process, which was maintained to almost normal TMJ structure, were expected to preserve the gap between the ramus and the zygomatic arch. CONCLUSION: Surgical intervention in the appropriate growth period is important to prevent the sequelae of mandibular growth suppression due to pediatric TMJ ankylosis.

Cost-Effectiveness Analysis of Perioperative Oral Management after Cancer Surgery and an Examination of the Reduction in Medical Costs Thereafter: A Multicenter Study.

In April 2012, perioperative oral management (POM) was approved for inclusion in the national health insurance system of Japan to prevent the occurrence of pneumonia, a major complication in cancer patients. The subsequent decrease in the incidence of postoperative pneumonia indicated the prophylactic effect of POM. The constant increase in health expenditure necessitates a cost-effectiveness analysis. In addition, the effect of reducing healthcare costs owing to health technologies must be evaluated. In the present multi-institutional study, the cost-effectiveness analysis of POM was conducted by comparing the incidence of postoperative pneumonia and the healthcare costs between patients who received surgery for malignant tumors before (n = 11,886) and after (n = 13,668) the introduction of POM. Additionally, the effect of reducing healthcare costs was evaluated. Reductions in the number of patients who developed pneumonia, duration of hospitalization, and number of deaths were observed after the introduction of POM. The incremental cost-effectiveness ratio was 111,927 yen, hence the prevention of postoperative pneumonia needs 111,927 yen per patient in healthcare costs. Consequently, a maximum reduction of 250,368,129 yen in healthcare costs was observed between the incremental costs for pneumonia treatment and the cost of POM. These findings indicate that improvements in cost-effectiveness can be expected in the future through the development of procedure and system for POM.

Preventive Effects of Sustainable and Developmental Perioperative Oral Management Using the "Oral Triage" System on Postoperative Pneumonia after Cancer Surgery.

Perioperative oral management is widely recognized in the healthcare system of Japan. Conventionally, the surgeon refers patients with oral problems to a dental or oral surgery clinic in the hospital. However, frequent in-house referrals were found to increase the number of incoming patients resulting in unsustainable situations due to an insufficient workforce. In 2011, the Center for Perioperative Medicine was established at our hospital to function as a management gateway for patients scheduled to undergo surgery under general anesthesia. The "oral triage" system, wherein a dental hygienist conducts an oral screening to select patients who need preoperative oral hygiene and functional management, was established in 2012. A total of 37,557 patients who underwent surgery at our hospital from April 2010 to March 2019 (two years before and seven years after introducing the system) were evaluated in this study. The sustainability and effectiveness of introducing the system were examined in 7715 cancer surgery patients. An oral management intervention rate of 20% and a significant decrease in the incidence of postoperative pneumonia (aOR = 0.50, p = 0.03) indicated that this system could be useful as a sustainable and developmental oral management strategy to manage surgical patients with minimal human resources.

Polo-like kinase in trypanosomes: an odd member out of the Polo family.

Polo-like kinases (Plks) are evolutionarily conserved serine/threonine protein kinases playing crucial roles during multiple stages of mitosis and cytokinesis in yeast and animals. Plks are characterized by a unique Polo-box domain, which plays regulatory roles in controlling Plk activation, interacting with substrates and targeting Plk to specific subcellular locations. Plk activity and protein abundance are subject to temporal and spatial control through transcription, phosphorylation and proteolysis. In the early branching protists, Plk orthologues are present in some taxa, such as kinetoplastids and Giardia, but are lost in apicomplexans, such as Plasmodium. Works from characterizing a Plk orthologue in Trypanosoma brucei, a kinetoplastid protozoan, discover its essential roles in regulating the inheritance of flagellum-associated cytoskeleton and the initiation of cytokinesis, but not any stage of mitosis. These studies reveal evolutionarily conserved and species-specific features in the control of Plk activation, substrate recognition and protein abundance, and suggest the divergence of Plk function and regulation for specialized needs in this flagellated unicellular eukaryote.

Pneumonia prevention effects of perioperative oral management in approximately 25,000 patients following cancer surgery.

AIM: We conducted a multicenter study to explore the risk factors of developing pneumonia and the effectiveness of perioperative oral management (POM) for the prevention of pneumonia in postsurgical patients. METHODS AND RESULTS: A survey covering eight regional hospitals was conducted over 4 years, from April 2010 to March 2014. Using the Diagnosis Procedure Combination database, a target group of 25,554 patients with cancer who underwent surgery was selected and assessed from a population of 346,563 patients without pneumonia on admission (sample population). The study compared the incidence of pneumonia and attempted to identify the significant predictive factors for its occurrence in these patients using multiple logistic regression analysis. Comparative assessment for the occurrence of pneumonia before and after POM implementation showed a significant incidence decrease after POM introduction in the target group, with no such change observed in the sample population. Multiple logistic regression analysis showed that the odds ratio for pneumonia occurrence after POM introduction was 0.44, indicating a reduced risk of pneumonia. CONCLUSION: POM in cancer patients was indeed effective in reducing the incidence of pneumonia in hospitals and thereby helped in preventing pneumonia during hospitalization.

The trypanosome-specific proteins FPRC and CIF4 regulate cytokinesis initiation by recruiting CIF1 to the cytokinesis initiation site.

The evolutionarily early divergent human parasite Trypanosoma brucei proliferates through binary cell fission in both its tsetse fly vector and mammalian host. The parasite divides unidirectionally along the longitudinal cell axis from the anterior cell tip toward the posterior cell tip through a mechanism distinct from that in the cells of its human host. Initiation of cytokinesis in T. brucei is regulated by two evolutionarily conserved protein kinases, the Polo-like kinase TbPLK and the Aurora B kinase TbAUK1, and a cohort of trypanosome-specific proteins, including the three cytokinesis initiation factors CIF1, CIF2, and CIF3. Here, using RNAi, in situ epitope tagging of proteins, GST pulldown, and coimmunoprecipitation assays, and immunofluorescence and scanning electron microscopy analyses, we report the identification and functional characterization of two trypanosome-specific proteins, flagellum attachment zone tip-localizing protein required for cytokinesis (FPRC) and CIF4. We found that the two proteins colocalize to the distal tips of the new and the old flagellum attachment zones and are required for cytokinesis initiation. Knockdown of FPRC or CIF4 disrupted the localization of CIF1, suggesting that they function upstream of CIF1. Moreover, depletion of CIF4 abolished FPRC localization, indicating that CIF4 acts upstream of FPRC. Together, these results identify two new cytokinesis regulators in T. brucei and integrate them into the CIF1-mediated cytokinesis regulatory pathway. These findings highlight the existence of a cytokinesis pathway in T. brucei that is different from that of its mammalian host and therefore suggest that cytokinesis in T. brucei could potentially be exploited as a new drug target.

A kinetochore-based ATM/ATR-independent DNA damage checkpoint maintains genomic integrity in trypanosomes.

DNA damage-induced cell cycle checkpoints serve as surveillance mechanisms to maintain genomic stability, and are regulated by ATM/ATR-mediated signaling pathways that are conserved from yeast to humans. Trypanosoma brucei, an early divergent microbial eukaryote, lacks key components of the conventional DNA damage-induced G2/M cell cycle checkpoint and the spindle assembly checkpoint, and nothing is known about how T. brucei controls its cell cycle checkpoints. Here we discover a kinetochore-based, DNA damage-induced metaphase checkpoint in T. brucei. MMS-induced DNA damage triggers a metaphase arrest by modulating the abundance of the outer kinetochore protein KKIP5 in an Aurora B kinase- and kinetochore-dependent, but ATM/ATR-independent manner. Overexpression of KKIP5 arrests cells at metaphase through stabilizing the mitotic cyclin CYC6 and the cohesin subunit SCC1, mimicking DNA damage-induced metaphase arrest, whereas depletion of KKIP5 alleviates the DNA damage-induced metaphase arrest and causes chromosome mis-segregation and aneuploidy. These findings suggest that trypanosomes employ a novel DNA damage-induced metaphase checkpoint to maintain genomic integrity.