Mck1-mediated proteolysis of CENP-A prevents mislocalization of CENP-A for chromosomal stability in Saccharomyces cerevisiae.

Centromeric localization of evolutionarily conserved CENP-A (Cse4 in Saccharomyces cerevisiae) is essential for chromosomal stability. Mislocalization of overexpressed CENP-A to non-centromeric regions contributes to chromosomal instability (CIN) in yeasts, flies, and humans. Overexpression and mislocalization of CENP-A observed in many cancers is associated with poor prognosis. Previous studies have shown that F-box proteins, Cdc4 and Met30 of the Skp, Cullin, F-box (SCF) ubiquitin ligase cooperatively regulate proteolysis of Cse4 to prevent Cse4 mislocalization and CIN under normal physiological conditions. Mck1-mediated phosphorylation of SCF-Cdc4 substrates such as Cdc6 and Rcn1 enhances the interaction of the substrates with Cdc4. Here, we report that Mck1 interacts with Cse4, and Mck1-mediated proteolysis of Cse4 prevents Cse4 mislocalization for chromosomal stability. Our results showed that mck1Δ strain overexpressing CSE4 (GAL-CSE4) exhibits lethality, defects in ubiquitin-mediated proteolysis of Cse4, mislocalization of Cse4 and reduced Cse4-Cdc4 interaction. Strain expressing GAL-cse4-3A with mutations in three potential Mck1 phosphorylation consensus site (S10, S16, and T166) also exhibits growth defects, increased stability with mislocalization of Cse4-3A, CIN, and reduced interaction with Cdc4. Constitutive expression of histone H3 (Δ16H3) suppresses the CIN phenotype of GAL-cse4-3A strain, suggesting that the CIN phenotype is linked to Cse4-3A mislocalization. We conclude that Mck1 and its three potential phosphorylation sites on Cse4 promote Cse4-Cdc4 interaction and this contributes to ubiquitin-mediated proteolysis of Cse4 preventing its mislocalization and CIN. These studies advance our understanding of pathways that regulate cellular levels of CENP-A to prevent mislocalization of CENP-A in human cancers.

DNAJC9 prevents CENP-A mislocalization and chromosomal instability by maintaining the fidelity of histone supply chains.

The centromeric histone H3 variant CENP-A is overexpressed in many cancers. The mislocalization of CENP-A to noncentromeric regions contributes to chromosomal instability (CIN), a hallmark of cancer. However, pathways that promote or prevent CENP-A mislocalization remain poorly defined. Here, we performed a genome-wide RNAi screen for regulators of CENP-A localization which identified DNAJC9, a J-domain protein implicated in histone H3-H4 protein folding, as a factor restricting CENP-A mislocalization. Cells lacking DNAJC9 exhibit mislocalization of CENP-A throughout the genome, and CIN phenotypes. Global interactome analysis showed that DNAJC9 depletion promotes the interaction of CENP-A with the DNA-replication-associated histone chaperone MCM2. CENP-A mislocalization upon DNAJC9 depletion was dependent on MCM2, defining MCM2 as a driver of CENP-A deposition at ectopic sites when H3-H4 supply chains are disrupted. Cells depleted for histone H3.3, also exhibit CENP-A mislocalization. In summary, we have defined novel factors that prevent mislocalization of CENP-A, and demonstrated that the integrity of H3-H4 supply chains regulated by histone chaperones such as DNAJC9 restrict CENP-A mislocalization and CIN.

Mad honey poisoning presenting with syncopal attack: a case report.

Mad honey contains grayanotoxin, which is commonly derived from the nectar of a few Rhododendron species. It is commonly used by natives of the Himalayas in the belief of its medicinal use. Case presentation: The authors report a case of 62 years old male with mad honey poisoning who was presented to the emergency department with loss of consciousness and had bradycardia and hypotension on arrival. The patient received intravenous fluids, atropine, and vasopressor support and was closely monitored in the coronary care unit for 48 h. Discussion: Grayanotoxin I and II are believed to be primarily responsible for mad honey intoxication and act by persistent activation of voltage-gated sodium channels. Hypotension, dizziness, nausea, vomiting, and impaired consciousness are the common presentation of mad honey intoxication. Toxic effects are usually mild and close monitoring for 24-48 h is sufficient but life-threatening complications like cardiac asystole, convulsions, and myocardial infarction have also been reported. Conclusion: Most cases of mad honey intoxication only need symptomatic treatment and close observation but the potential for deterioration and life-threatening complications must also be considered.

The histone H3/H4 chaperone CHAF1B prevents the mislocalization of CENP-A for chromosomal stability.

Restricting the localization of the evolutionarily conserved centromeric histone H3 variant CENP-A to centromeres prevents chromosomal instability (CIN). The mislocalization of CENP-A to non-centromeric regions contributes to CIN in yeasts, flies and human cells. Even though overexpression and mislocalization of CENP-A have been reported in cancers, the mechanisms responsible for its mislocalization remain poorly understood. Here, we used an imaging-based high-throughput RNAi screen to identify factors that prevent mislocalization of overexpressed YFP-tagged CENP-A (YFP-CENP-A) in HeLa cells. Among the top five candidates in the screen - the depletion of which showed increased nuclear YFP-CENP-A fluorescence - were the histone chaperones CHAF1B (or p60) and CHAF1A (or p150). Follow-up validation and characterization experiments showed that CHAF1B-depleted cells exhibited CENP-A mislocalization, CIN phenotypes and increased enrichment of CENP-A in chromatin fractions. The depletion of DAXX, a histone H3.3 chaperone, suppressed CENP-A mislocalization and CIN in CHAF1B-depleted cells. We propose that in CHAF1B-depleted cells, DAXX promotes mislocalization of the overexpressed CENP-A to non-centromeric regions, resulting in CIN. In summary, we identified regulators of CENP-A localization and defined a role for CHAF1B in preventing DAXX-dependent CENP-A mislocalization and CIN.

Assessment of App Store Description and Privacy Policy to Explore Ethical and Safety Concerns Associated with the Use of Mental Health Apps for Depression.

Background: Mental Health apps (MH apps) could help address the huge unmet mental health care need of developing countries. This study aimed to explore potential ethical, data safety, and privacy issues associated with using MH apps for depression. Methods: A cross-sectional assessment of the top 50 MH apps (by Google Play store search result ordering) for depression available in India was conducted in November 2021. Results: Most apps were listed under the category of health and fitness (54%). The median number of total and dangerous permissions requested at the time of download was nine and three, respectively. Privacy policy in English was available for 76%. The average length of the privacy policy was 2171 words, and the mean Flesch-Kincaid reading grade level was 12 (much higher than the recommended cut-off of eight). Important features relevant to safeguarding consumer confidentiality, including names of third parties with which user data could be shared (42%), explicit consent before sharing data with third parties (16%), and assurance regarding the collection of de-identifiable data (11%), were missing from the majority of privacy policies. Conclusion: There is an urgent need to improve the accessibility and usability of privacy policies by app developers, with the active involvement of other stakeholders.

Matrin3 regulates mitotic spindle dynamics by controlling alternative splicing of CDC14B.

Matrin3 is an RNA-binding protein that regulates diverse RNA-related processes, including mRNA splicing. Although Matrin3 has been intensively studied in neurodegenerative diseases, its function in cancer remains unclear. Here, we report Matrin3-mediated regulation of mitotic spindle dynamics in colorectal cancer (CRC) cells. We comprehensively identified RNAs bound and regulated by Matrin3 in CRC cells and focused on CDC14B, one of the top Matrin3 targets. Matrin3 knockdown results in increased inclusion of an exon containing a premature termination codon in the CDC14B transcript and simultaneous down-regulation of the standard CDC14B transcript. Knockdown of CDC14B phenocopies the defects in mitotic spindle dynamics upon Matrin3 knockdown, and the elongated and misoriented mitotic spindle observed upon Matrin3 knockdown are rescued upon overexpression of CDC14B, suggesting that CDC14B is a key downstream effector of Matrin3. Collectively, these data reveal a role for the Matrin3/CDC14B axis in control of mitotic spindle dynamics.

Context-Dependent Function of Long Noncoding RNA PURPL in Transcriptome Regulation during p53 Activation.

PURPL is a p53-induced lncRNA that suppresses basal p53 levels. Here, we investigated PURPL upon p53 activation in liver cancer cells, where it is expressed at significantly higher levels than other cell types. Using isoform sequencing, we discovered novel PURPL transcripts that have a retained intron and/or previously unannotated exons. To determine PURPL function upon p53 activation, we performed transcriptome sequencing (RNA-Seq) after depleting PURPL using CRISPR interference (CRISPRi), followed by Nutlin treatment to induce p53. Strikingly, although loss of PURPL in untreated cells altered the expression of only 7 genes, loss of PURPL resulted in altered expression of ~800 genes upon p53 activation, revealing a context-dependent function of PURPL. Pathway analysis suggested that PURPL is important for fine-tuning the expression of specific genes required for mitosis. Consistent with these results, we observed a significant decrease in the percentage of mitotic cells upon PURPL depletion. Collectively, these data identify novel transcripts from the PURPL locus and suggest that PURPL delicately moderates the expression of mitotic genes in the context of p53 activation to control cell cycle arrest.

Pregnancy with heart disease in South Asia: A systematic review and meta-analysis of prevalence and outcome.

Background: The prevalence and paradigm of antenatal heart disease are contrasting between high-income, industrialized, and low- and middle-income countries. In this systematic review, we report the prevalence of heart disease and its spectrum in pregnant women of South Asia. Methods: We searched through different electronic databases (PubMed, Google-scholar, Embase, Cochrane Library) to locate relevant articles. Studies with sufficient data that met our inclusion criteria were included. Two reviewers independently screened the articles. Discrepancies were resolved by other reviewers. Subsequently, data extraction was done using a standardized form and quality assessment of studies using the Joanna Briggs Institute tool. Meta-analysis was done using R software. Results: After various stages of screening 25 studies were included in the final quantitative synthesis. The pooled prevalence of heart disease among pregnant women was 1.46% (95% CI 0.99-2.01). Among those with heart disease, 70.25% (95% CI 64.87-75.38) had Rheumatic heart disease and 18.10% (95% CI 14.39-22.12) had congenital heart disease. The pooled prevalence of preterm labor and delivery among pregnant women with heart disease was 17.63% (95% CI 12.18-23.80). Similarly, the pooled maternal and fetal mortality rates were 26.14 (95% CI 12.47-43.55) and 50.48 (95% CI 29.59-75.83) per 1000 pregnant women with heart disease respectively. Conclusion: As pregnancy, itself is a prolonged state of physiologic stress, heart disease further adds to the risk both for the mother and fetus. Having such a high prevalence, efforts must be made to detect and closely monitor the condition antenatally, and decisions should be made according to the clinical conditions of the patient.

Interaction of Phthalates with Lipid Bilayer Membranes.

Phthalates are esters of phthalic acid, widely used as additives in the manufacture of plastics. They are not covalently linked to polymer chains and can easily leach out, disperse in the environment, and get into contact with living organisms. Several short chain phthalates are classified as endocrine disruptors or hormonal active agents, and have also been reported to promote various kinds of cancer. However, the biological effects of longer chain analogues are less well known. Moreover, little is known on the permeation of phthalates and their metabolites through biological membranes and on their effects on the physical properties of membranes. Here we explore the interaction of a group of phthalates and their main metabolites with model biological membranes. We focus on three industrially relevant phthalates, with acyl chains of different sizes, and their monoester metabolites. We use molecular dynamics simulations to predict the distribution in model membranes, as well as permeabilities and effects on the structural, dynamic, and elastic properties of the membranes. We find that alterations of membrane properties are significant and only weakly affected by the size of acyl chains, suggesting that modifications of molecular size may not be sufficient to reduce the impact of this class of molecules on the environment and health.

Intravenous Ringers lactate versus normal saline for predominantly mild acute pancreatitis in a Nepalese Tertiary Hospital.

BACKGROUND: Acute pancreatitis (AP) is a common presentation in patients admitted with acute abdomen. Whether Ringers lactate (RL) or Normal Saline (NS) as a resuscitation fluid is better still remains unclear. The aim of this study is to compare the efficacy of RL and NS in terms of control of systemic inflammation by measuring indirect markers specifically Systemic Inflammation Response Syndrome (SIRS) scores and C- Reactive Protein (CRP) level. METHODS: This was an open label randomized trial conducted in a tertiary level hospital of Nepal. Ethical approval was obtained prior to the study. Patients with acute pancreatitis were randomized to either RL or NS group for the fluid resuscitation. The fluid was given as per the study protocol for three days for hydration. Baseline SIRS and CRP were recorded on admission and subsequently as defined. All the data were analyzed using SPSS ver 20.0 software. RESULTS: Total 51 patients were enrolled, 26 in RL and 25 in NS group. The commonest etiology of AP was alcohol (84.31%). SIRS was present in 46.2% and 64.0% of patients in RL and NS group respectively (p = 0.20) on admission. At least one SIRS criteria was still present in 44.0% of patients in the NS group compared to only 15.4% in the RL group after 24 hours (p = 0.025). The baseline CRP were comparable in both the groups. However after 72 hours, the increment of CRP was more in the NS group compared to the RL group; median value of 14.2 mg/dl (12.15, 16.45) and 22.2 mg/dl (18.20, 30.60) in RL and NS group respectively (p<0.001). CONCLUSIONS: Ringers lactate was associated with a reduction in systemic inflammation compared to normal saline in patients with acute pancreatitis. Incidence of SIRS at 72 hours and occurrence of local complications were however similar in both the groups.

Counteraction between Astrin-PP1 and Cyclin-B-CDK1 pathways protects chromosome-microtubule attachments independent of biorientation.

Defects in chromosome-microtubule attachment can cause chromosomal instability (CIN), frequently associated with infertility and aggressive cancers. Chromosome-microtubule attachment is mediated by a large macromolecular structure, the kinetochore. Sister kinetochores of each chromosome are pulled by microtubules from opposing spindle-poles, a state called biorientation which prevents chromosome missegregation. Kinetochore-microtubule attachments that lack the opposing-pull are detached by Aurora-B/Ipl1. It is unclear how mono-oriented attachments that precede biorientation are spared despite the lack of opposing-pull. Using an RNAi-screen, we uncover a unique role for the Astrin-SKAP complex in protecting mono-oriented attachments. We provide evidence of domains in the microtubule-end associated protein that sense changes specific to end-on kinetochore-microtubule attachments and assemble an outer-kinetochore crescent to stabilise attachments. We find that Astrin-PP1 and Cyclin-B-CDK1 pathways counteract each other to preserve mono-oriented attachments. Thus, CIN prevention pathways are not only surveying attachment defects but also actively recognising and stabilising mature attachments independent of biorientation.

CENP-A overexpression promotes aneuploidy with karyotypic heterogeneity.

Chromosomal instability (CIN) is a hallmark of many cancers. Restricting the localization of centromeric histone H3 variant CENP-A to centromeres prevents CIN. CENP-A overexpression (OE) and mislocalization have been observed in cancers and correlate with poor prognosis; however, the molecular consequences of CENP-A OE on CIN and aneuploidy have not been defined. Here, we show that CENP-A OE leads to its mislocalization and CIN with lagging chromosomes and micronuclei in pseudodiploid DLD1 cells and xenograft mouse model. CIN is due to reduced localization of proteins to the kinetochore, resulting in defects in kinetochore integrity and unstable kinetochore-microtubule attachments. CENP-A OE contributes to reduced expression of cell adhesion genes and higher invasion of DLD1 cells. We show that CENP-A OE contributes to aneuploidy with karyotypic heterogeneity in human cells and xenograft mouse model. In summary, our results provide a molecular link between CENP-A OE and aneuploidy, and suggest that karyotypic heterogeneity may contribute to the aggressive phenotype of CENP-A-overexpressing cancers.

Sacrococcygeal Teratoma: A Case Report.

Sacrococcygeal teratoma is rare and happens in 1:35,000 to 40,000 live births. It is more common in girls than boys with the reported ratio of 3:1 to 4:1. We herein report an unusual case of a huge sacrococcygeal teratoma, which was more than half of the size and weight of the baby which was terminated at 24 weeks of gestation.

Development of IglC and GroEL recombinant vaccines for francisellosis in Nile tilapia, Oreochromis niloticus.

Warm-water piscine francisellosis is a granulomatous bacterial disease caused by Francisella orientalis (Fo). The disease has been detected in a wide range of fish species globally, causing mortalities as high as 90% and significant economic losses. Currently there are no commercially available vaccines and few treatment options exist. In the current study, two novel recombinant vaccines were prepared using diatom-expressed IglC or bacterial-expressed GroEL proteins. The vaccine antigens were emulsified with either nanoparticles or a commercially available oil-based adjuvant. Nile tilapia, Oreochromis niloticus, fingerlings were immunized intracoelomically with the recombinant IglC or GroEL vaccines, diatoms alone or phosphate buffer saline. Approximately 840-degree days post-vaccination, fish were challenged via immersion with 106 CFU/mL of wild-type Fo. Twenty-one days post challenge (dpc), the highest relative percent survival was recorded in the IglC-Montanide group (75%), compared to 53%, 50%, 22%, 19% and 16% in the IglC-nanoparticles, GroEL-Montanide, GroEL-nanoparticles, diatoms-Montanide and diatoms-nanoparticles groups, respectively. Protection correlated with significantly higher specific antibody responses in the IglC-Montanide group. Moreover, a significantly lower bacterial load was detected in spleen samples from the IglC-Montanide survivor tilapia compared to the other experimental groups. This is the first report of recombinant vaccines against piscine francisellosis in tilapia. The Fo vaccines described in our study may facilitate development of a safe, cost-effective and highly protective vaccine against francisellosis in farmed tilapia.

A randomized, placebo-controlled, phase II study of obeticholic acid for primary sclerosing cholangitis.

BACKGROUND & AIMS: Primary sclerosing cholangitis (PSC) is a rare, cholestatic liver disease with no currently approved therapies. Obeticholic acid (OCA) is a potent farnesoid X receptor (FXR) agonist approved for the treatment of primary biliary cholangitis. We investigated the efficacy and safety of OCA in patients with PSC. METHODS: AESOP was a phase II, randomized, double-blind, placebo-controlled, dose-finding study. Eligible patients were 18 to 75 years of age with a diagnosis of PSC and serum alkaline phosphatase (ALP) ≥2× the upper limit of normal (ULN) and total bilirubin <2.5× ULN. Patients were randomized 1:1:1 to receive placebo, OCA 1.5-3.0 mg, or OCA 5-10 mg once daily for a 24-week, double-blind phase followed by a 2-year, long-term safety extension (LTSE). Primary endpoints were change in ALP from baseline to week 24, and safety. RESULTS: The intent-to-treat population comprised 76 patients randomized to placebo (n = 25), OCA 1.5-3.0 mg (n = 25), and OCA 5-10 mg (n = 26). At week 24, serum ALP was significantly reduced with OCA 5-10 mg vs. placebo (least-square [LS] mean difference = -83.4 [SE = 40.3] U/L; 95% CI -164.28 to -2.57; p = 0.043). Serum ALP was not significantly reduced with OCA 1.5-3.0 mg vs. placebo at week 24 (LS mean [SE] difference = -78.29 [41.81] U/L; 95% CI -162.08 to 5.50; p = 0.067). Total bilirubin remained comparable to baseline in all groups. The most common treatment-emergent adverse event was dose-related pruritus (placebo 46%; OCA 1.5-3.0 mg 60%; OCA 5-10 mg 67%). Reductions in ALP were maintained during the LTSE, and no new safety signals emerged. CONCLUSIONS: Treatment with OCA 5-10 mg reduced serum ALP in patients with PSC. Mild to moderate dose-related pruritus was the most common adverse event. REGISTRATION: ClinicalTrials.gov: NCT02177136; EudraCT: 2014-002205-38. LAY SUMMARY: Primary sclerosing cholangitis (PSC) is a long-term disease that damages the bile ducts in the liver over time. In the AESOP clinical study in patients with PSC, obeticholic acid reduced serum alkaline phosphatase (a potential marker of disease severity) during an initial 24-week treatment period. The result was sustained during the 2-year, long-term extension of the study. The most common side effect of obeticholic acid in the study was itchy skin, which is consistent with earlier clinical studies.

MARK2/Par1b kinase present at centrosomes and retraction fibres corrects spindle off-centring induced by actin disassembly.

Tissue maintenance and development requires a directed plane of cell division. While it is clear that the division plane can be determined by retraction fibres that guide spindle movements, the precise molecular components of retraction fibres that control spindle movements remain unclear. We report MARK2/Par1b kinase as a novel component of actin-rich retraction fibres. A kinase-dead mutant of MARK2 reveals MARK2's ability to monitor subcellular actin status during interphase. During mitosis, MARK2's localization at actin-rich retraction fibres, but not the rest of the cortical membrane or centrosome, is dependent on its activity, highlighting a specialized spatial regulation of MARK2. By subtly perturbing the actin cytoskeleton, we reveal MARK2's role in correcting mitotic spindle off-centring induced by actin disassembly. We propose that MARK2 provides a molecular framework to integrate cortical signals and cytoskeletal changes in mitosis and interphase.

End-hole Versus Microvalve Infusion Catheters in Patients Undergoing Drug-Eluting Microspheres-TACE for Solitary Hepatocellular Carcinoma Tumors: A Retrospective Analysis.

INTRODUCTION: Pre-transplant locoregional therapy for hepatocellular carcinoma (HCC) during bridge-to-transplant impacts recurrence and survival rates following liver transplantation. Optimizing the effectiveness of transarterial chemoembolization (TACE) in this population is imperative, and microvalve infusion catheters offer a means of such improvement. METHODS: All treatment-naive patients with solitary HCC tumors < 6.5 cm who underwent drug-eluting microspheres (DEM) TACE between 04/2015 and 08/2017 were retrospectively reviewed. Eighty-eight included patients underwent DEM-TACE with either standard end-hole catheters (EH) or microvalve infusion catheters (MVI). The EH (n = 70) and MVI (n = 18) cohorts had similar baseline tumor size, laboratory values, and tumor etiologies. RESULTS: Initial objective response rates were significantly higher in MVI vs. EH (100% vs. 76.5%, p = 0.019). There was no difference in adverse events between groups (p = 0.265). MVI patients exhibited lower AST (p = 0.003) and ALT (p = 0.044) at 6 months. Blinded pathological analysis of explanted livers showed greater concentrations of microspheres within the tumor relative to the surrounding tissue in MVI explants (88.7 ± 10.6%) versus the EH explants (55.3 ± 32.7%) (p = 0.002). There was significantly higher percentage tumor necrosis in the MVI group (89.0 ± 2.2%) compared with the EH group (56.1 ± 44.5%) (p = 0.006). CONCLUSION: In this retrospective study of a single-center cohort, DEM-TACE procedures with MVI were associated with improved tumor response, increased deposition of microspheres within tumor tissue, and higher percentage tumor necrosis at explant relative to those performed using EH catheters.

Spindle rotation in human cells is reliant on a MARK2-mediated equatorial spindle-centering mechanism.

The plane of cell division is defined by the final position of the mitotic spindle. The spindle is pulled and rotated to the correct position by cortical dynein. However, it is unclear how the spindle's rotational center is maintained and what the consequences of an equatorially off centered spindle are in human cells. We analyzed spindle movements in 100s of cells exposed to protein depletions or drug treatments and uncovered a novel role for MARK2 in maintaining the spindle at the cell's geometric center. Following MARK2 depletion, spindles glide along the cell cortex, leading to a failure in identifying the correct division plane. Surprisingly, spindle off centering in MARK2-depleted cells is not caused by excessive pull by dynein. We show that MARK2 modulates mitotic microtubule growth and length and that codepleting mitotic centromere-associated protein (MCAK), a microtubule destabilizer, rescues spindle off centering in MARK2-depleted cells. Thus, we provide the first insight into a spindle-centering mechanism needed for proper spindle rotation and, in turn, the correct division plane in human cells.

Development of a silicon limitation inducible expression system for recombinant protein production in the centric diatoms Thalassiosira pseudonana and Cyclotella cryptica.

BACKGROUND: An inducible promoter for recombinant protein expression provides substantial benefits because under induction conditions cellular energy and metabolic capability can be directed into protein synthesis. The most widely used inducible promoter for diatoms is for nitrate reductase, however, nitrogen metabolism is tied into diverse aspects of cellular function, and the induction response is not necessarily robust. Silicon limitation offers a means to eliminate energy and metabolic flux into cell division processes, with little other detrimental effect on cellular function, and a protein expression system that works under those conditions could be advantageous. RESULTS: In this study, we evaluate a number of promoters for recombinant protein expression induced by silicon limitation and repressed by the presence of silicon in the diatoms Thalassiosira pseudonana and Cyclotella cryptica. In addition to silicon limitation, we describe additional strategies to elevate recombinant protein expression level, including inclusion of the 5' fragment of the coding region of the native gene and reducing carbon flow into ancillary processes of pigment synthesis and formation of photosynthetic storage products. We achieved yields of eGFP to 1.8% of total soluble protein in C. cryptica, which is about 3.6-fold higher than that obtained with chloroplast expression and ninefold higher than nuclear expression in another well-established algal system. CONCLUSIONS: Our studies demonstrate that the combination of inducible promoter and other strategies can result in robust expression of recombinant protein in a nuclear-based expression system in diatoms under silicon limited conditions, separating the protein expression regime from growth processes and improving overall recombinant protein yields.